172 terms


Verbage from Fong's syllabus
Growth factor receptor signaling pathway
Type of tyrosine kinase signaling pathway. Can be problematic when activated (as this can cause cancer).
1. Can phosphorylate themselves and other cytoplasmic proteins at their tyrosine residues.
2. Binding of growth factors/ligands results in dimerization or clustering of the receptors, which then allows autophosphorylation.
3. The phosphorylated receptor acts through one or more conserved non-catalytic domains, which trigger intracellular cascades.
Fibroblast growth factor receptors
1. Transmembrane domain.
2. Extracellular: 3 immunoglobulin-like domains. Form loop-structures held together by an intra-chain disulfide bonds between cysteine residues.
3. Intracellular: Split tyrosine kinase domain.
Signaling pathway:
1. When FGF's are in close proximity to cell surface, bind heparan sulfate proteoglycan (through heparan sulfate binding domain of FGF to S-domain of heparan sulfate).
2. S-domain binds to lysine rich region of 2nd immuoglobulin-like domain of FGFR.
3. 2 combined with FGF's binding to FGFR's leads to dimerization of FGFR's and thus autophosphorylation.
4. This begins an intracellular cascade (differs depending on tissue type).
Activation mutations:
1. Increased binding affinity of the ligand.
2. RECEPTOR DIMERIZATION DEFECTS: Increased propensity of receptors to dimerize. One example is, when cysteine residues are lost in the immuoglobulin-like domains of FGFR's, inter-chain disulfide bonds are formed instead of intra-chain disulfide bonds.
1. Crouzon Syndrome - FGFR2 - Autosomal dominent, some non-penetrance and variable expression. Variable craniosynostosis, ocular proptosis due to shallow orbits, mid-face hypoplasia, conductive hearing loss, some mental deficiency.
2. Apert Syndrome - FGFR2 - Craniosynostosis, Total syndactyly of the fingers and toes, congential heart disease.
3. Pfeiffer syndrome - FGFR2 or FGFR1 - Craniosynostosis, prominent thumbs and big toes.
4. Muenke syndrome - FGFR3 - Variable unilateral coronal synostosis (generally some other findings).
5. Achondroplasia - FGFR3 (transmembrane) - Autosomal dominant 80% new mutations all of paternal origin - Problem with long bone formation. Short stature, small chest, macrocrania with small foramen magnum and midface hypoplasia. Normal intelligence. Narrow airways possible.
6. Thanatophoric Dysplasia - FGFR3 - More severe problem with long bone formation. Generally result in stillbirths or early infant deaths secondary to lung hypoplasia. Bones are extremely short and disfigured. All cases are sporadic bc no one reproduces.
7. Hypochondroplasia - FGFR3 - Milder problem with long bone formation. Individuals are usually taller and there is a lot of variability.
Secondary craniosynostosis
When brain growth suddenly stops, then osteoblast activity outpaces brain growth and thus sutures will close.
Primary craniosynostosis
Normally, during fetal growth, brain growth stretches the dura mater, which responds by releasing fibroblast growth factors, which in turn increase osteoblast activity on the skull. Thus, as the skull is separated, it grows.
However, when Fibroblast growth factor receptors (FGFR's) are extra active, then effectively unregulated osteoblast activity can close the sutures early.
Often, not all the sutures close early which can cause different effects. For example, premature closure of:
1. Sagittal suture results in an elongated head shape with narrow bifrontal diameters. Called dolicocephaly or scaphocephaly.
2. Metopic suture (from midline eye to upper forhead) results in an anterior narrowing of the head. Called trigonocephaly.
3. Coronal or lambdoid sutures result in short anterior-posterior diameter of head, with some widening laterally. Called brachycephaly.
4. One coronal or lambdoid suture results in an asymmetric shortening of the head called plagiocephaly (this is primary, secondary plagiocephaly is generally a consequence of fetal deformation or sleep positions).
Histone modifications
General principle: DNA is negatively charged while histones are positively charged. So, to activate regions of DNA, you just need to decrease positive charge of histones (which will decrease the affinity). The reverse is true.
1. Phosphorylation: 1st step to destabilize a compacted chromatin region.
a. Histone H1 is phosphorylated at serine and threonine residues by H1 kinase enzymes. This results in adjacent H1's being repulsed from one another (and eventually dissociate from chromatin). H1 phosphatases reverse this process and help compact chromatin.
b. H1 proteins are often replaced by High mobility group proteins (HMG proteins) which are non-histone.
c. Phosphorylation of core histones occurs via MSK1 and MSK2 (mitogen/stress induced kinases 1 and 2) and RSK2 (ribosomal S6 kinase) which are under control of MAP kinase cascade. This serves to decrease positive charge and to promote interaction of histones with acetyl- and methyltransferases.
2. Acetylation: Major step that neutralizes positive histone charge through acetylation of lysine residues.
a. Acetyl group derived from an acetyl CoA is transferred to lysine via histone acetyltransferase (HAT) (activates chromatin).
b. HAT both provides its catalytic domain and as a scaffold protein through which other proteins can interact.
c. Deacetylation occurs via histone deacetylases (HDAC). One type of HDAC called Sir2-like histone deacetylase or sirtuins, leads to deacetylation through a NAD mediated process. This represses DNA.
3. Methylation:
a. Methylation at lysine or arginine residues occurs via histone methyltransferases with SAM (S-adenosylmethionine) serving as methyl group donor.
b. Can occur at a single residue multiple times.
c. Can lead to activation or deactivation. Some arginine or lysine residues in H3 need to be methylated for transcription activation, but others can silence transcription.
d. Histones can be demethylated via e-N-methyllysine demethylase which oxidizes methyl group to formaldehyde or Peptidyle arginine demethylase, which deaminates methylated arginine residues to citrulline residues.
4. Poly(ADP-ribosylation): Strongly negative charge so acts as an activator (decreases histone positive charge).
a. ADP-ribosyl residue is transferred from NAD to the carboxyl group of glutamate and aspartate residues of histones via poly(ADP-ribosyl) polymerase (PARP1).
b. Reverse reaction is via poly(ADP-ribose) glycohydrolase.
5. Ubiquitylation:
a. Mono-ubiquitylation at core or H1 proteins triggers histone methylation inactivation and silencing.
b. Therefore, E3 ubiquitin ligases, which remove ubiquitin groups from histones, are transcriptional co-activators.
ATP-dependent nucleosome remodeling
Chromatin remodeling is mainly done by ATP-dependent motor proteins like SWI/SNF complex (SWItching and Sucrose-Non-Fermenting).
Two primary movements:
1. Sliding - movement of nucleosome along DNA.
2. Structural alteration - Partial or complete disruption of nucleosome.
DNA methylation
Mainly happens at CpGs by action of methyltransferases.
CpG islands: Regions of DNA as long as 1kb in length that contain a high percentage of CpG dinucleotides. Most are around promoter regions of "house-keeping" genes aka genes that are constitutively active in most cells. These tend to be UNMETHYLATED and their methylation tends to result in gene silencing.
Outside of CpG islands: Most CpG dinucleotides are methylated and thus repressed (less access by transcription factors). Hyper-methylated in LI and Alu sequences.
Methylation in development:
1. Primordial germ cells, DNA methylation is low. While germ cells mature, methylation levels increase (especially in sperm).
2. After fertilization, dramatic decrease in methylation (since there is so much gene expression going on).
3. Blastocyst stage means more methylation, because cell differentiation means different genes are getting methylated for differentiation.
4. Trophoblast cells have very low levels of methylation, since low need for differentiation.
DNA methylation mediated repression:
1. DNA is methylated via methyltransferases.
2. Methylated DNA is recognized by methyl-CpG-binding proteins (MeCP) which recruit HDAC's and other chromatin modifying activities that accomplish repression.
3. Deficiency of MeCP2 results in Rett syndrome.
Epigenetic inheritance
DNA methylation is maintained from one cell generation to the next (think imprinting and X-inactivation). DNA methyltransferases will recognized hemimethylated DNA after replication (so, if start with CmethpG-GpCmeth and end with CmethpG-GC and CpG-GpCmeth, the unmethylated C's will be methylated by methyltransferases).
1. Immunodeficiency-centromeric instability-facial syndrome (ICF syndrome) - Caused by defect in DNA methyltransferase 3B (DNMT3B). Symptoms are minor facial anomalies, growth retardation, variable immunodeficiency and bizarre chromosomal rearrangements, especially involving chromosomes 1, 9, and 16 which have large satellite DNA that are normally methylated but aren't in this syndrome.
Heterochromatin is the portion of the genome that remains condensed throughout the cell cycle. Generally has histone hypoacetylation and methylation of H3K9. Forms via these steps:
1. Repressive factors nucleate at a silencer and modify nearby nucleosomes by deacetylation and/or methylation of H3K9.
2. Spreading: HP1 recognizes and binds to nucleosomes methylated at H3K9. HP1 then recruits more histone methyltransferase so the next set of nucleosomes is also methylated at H3K9. This can encompass an entire chromosome (X-inactivation).
Genomic imprinting
Some genomic regions contain alleles that are only active if inherited from the father and inactive only if inherited from the mother (or vice versa). Not determined by the DNA code.
Note that in the offspring gametes, all alleles are de-imprinted and then re-imprinted depending on the sex of the offspring. For example, if a male imprint thing gets inherited by a female, the female's gametes will de-imprint that allele such that it is de-imprinted when it is passed on.
Prader-Willi Syndrome (PWS)
Disease of genomic imprinting.
1. Generally related to a small deletion in same spot as Angleman syndrome that is always of paternal origin (AS is always maternal). This is because normally the region is maternally imprinted, so a paternal deletion leads to no gene products from the region.
2. Can also be caused by maternal uniparental disomy. This means they have two maternal alleles of the PWS region, which are both imprinted, so no gene products can be produced.
1. Neonatal hypotonia
2. Hyperphagia
3. Obesity
4. Hypogonadism
5. Developmental disabilities and dysmorphic features.
1. Methylation sensitive restriction enzymes are the most sensitive test.
Angleman Syndrome (AS)
Disease of genomic imprinting.
1. Generally related to small deletion in same spot as Prader-Willi Syndrome that is always of maternal origin (PWS is always paternal). This is because normally the region is paternally imprinted, so a maternal deletion leads to no gene products from the region.
2. Can also be caused by paternal uniparental disomy. This means they have two paternal alleles of the AS region, which are both imprinted, so no gene products can be produced.
1. Puppet-like hand movements.
2. Spontaneous unprovoked laughter.
3. Ataxic gait
4. Severe developmental delay including absent speech.
5. Seizure disorder
6. Hypopigmentation and characteristic facial features.
1. 80% (70% deletion related and 7% via uniparental disomy) have abnormal methylation patterns and so can be detected with methylation detecting restriction enzymes.
2. 3% have mutation in imprinting center. This can be passed on to offspring (the rest generally can't be).
3. 11% have a more traditional mutation destroying function of the gene in the imprinted region. Detected via FISH or CMA.
Uniparental disomy
When both alleles of a genetic locus in an individual are inherited from the same parent (has a role in Anglemane Syndrome and Prader-Willi Syndrome).
Two types:
1. Uniparental Isodisomy - When the 2 UPD alleles have the same grand-parental origin. Represents the duplication and transmission (or transmission and duplication) of a single parental chromosome.
2. Uniparental Heterodisomy - When the 2 UPD alleles have different grand-parental origins. Represents the transmission of both copies of the homologous chromosome pair from a single parent. Two mechanisms for this:
a. Fertilization of a nullisomic gamete by a disomic gamete, both of which are products of a meiotic non-disjunction.
b. Fertilization of a disomic gamete by a normal gamete to form a trisomic zygote, followed immediately by trisomy rescue due to anaphase lag.
Caused by mechanical constraint or pressure on the developing fetus resulting in distortion of the normal developmental process. Common causes (most are space constraints within uterus):
1. Multiple gestations
2. Oligohydramnios (not enough amniotic fluid)
3. Breech presentation
4. Maternal uterine anomalies
5. Uterine tumors
6. Inactivity due to neuromuscular abnormalities
7. Congenital anomalies resulting in space-occupying lesion (eg teratoma).
8. Large fetus secondary to genetic syndrome (maternal diabetes, Beckwith-Wiedemann syndrome).
1. Facial asymmetry or positional micrognathia (small jaw)
2. Plagiocephaly with or without torticollis (contracture of the sternocleidomastoid muscle)
3. Congenital hip dislocation
4. Joint contractures
5. Aplasia cutis (absense of skin) secondary to friction between fetal skin and the uterine wall (generally due to oligohydramnios).
Disruption of fetus
Interruption of normal developmental processes. For example, amnion rupture sequence is when the amnion ruptures and leads to the formation of small strands of amnion which can encircle developing fetal structures (eg limbs or face) resulting in annular constrictions, amputations, and facial clefting. Generally happens in 1st 12 weeks of gestation.
When a single defective gene is sufficient to cause the multiple manifestations of the genetic syndrome (which often involve multiple organ systems). Examples as to how this could happen:
1. Gene product plays a role in global or regional developmental processes such as axis determination, patterning, or segmentation.
2. Gene product plays a critical role in many different pathways of development.
3. Gene product plays a critical role in the development of one tissue or cell type which plays a role in the development of many organ systems (eg neural crest survival and migration).
Malformation Sequence
Often mistaken for pleiotropy. Multiple anomalies of the fetus are not the result of a pleiotropic gene effect, but that of a logical chain of events that begin with a seemingly unrelated genetic issue. Example of this is Oligohydramnios or Potter sequence:
1. Any renal anomaly that is associated with decreased fetal urine production results in decrease of amniotic fluid (oligohydramnios).
2. This leads to restriction of fetal limb movement, resulting in congenital limb contracture.
3. Facial deformation and skin defects secondary to the same deformational process are also common.
4. External compression from extreme oligohydramnios results in the under-development of the lungs.
Continuous gene syndrome
Often mistaken for pleiotropy. If there is a cluster of genes that are all linked to different congenital malformations, then deletions with varying break points can result in different constellations of malformations.
a. DiGeorge syndrome and Velocardiofacial syndrome (both from the deletions in the same continuous region). Detection via FISH. Symptoms:
1. Conotruncal congenital heart disease
2. palatal defect or insufficiency
3. Mandibular hypoplasia
4. Hypoparathyroidism
5. Immunodeficiency
6. Craniofacial defects
b. WAGR (Wilms tumor, aniridia, gentiourinary anomaly, and retardation) syndrome.
Large polymers comprised of tubulin and microtubule-associated proteins. These tend to extend the entire length of the cell.
1. Scaffolding
2. Spatial organization to organelles within the cell
3. Components of mitotic spindle, cilia, and flagella.
4. Major system for intracellular trafficking.
Large polymers comprised of actin and actin-binding proteins that localize to plasma membrane (space just below the cell cortex) and in microvilli.
1. Cell shape and structure (including during cell division and migration)
2. Intracellular trafficking.
Intermediate Filaments
Tissue specific filaments that have a variety of functions. Most commonly associated with nuclear membrane. Note: NOT intracellular trafficking.
RBC surface area/volume (and associated disorders)
Generally, surface area is a function of membrane-cytoskeleton interactions while volume is a function of membrane-associated ion transporters. RBC's can stand 250% deformation linearly but only 3-4% increase in surface area. This is why they are shaped like biconcave discs - they have more reserve surface area for going through narrow capillaries.
RBC membrane:
1. Composed of cholesterol and phospholipid bilayer. Cholesterol is evenly distributed between the two layers, but phospholipids are asymmetrically arranged by proteins called flippases, floppases, or scramblases.
2. Spectrin - Helps with shape and flexibility of RBC membrane due to formation of a fishing-net structure created by spectrin interacting with short actin filaments (14-16 subunits in length). Composed of triple-helix alpha and beta subunits connected anti-parellelly. Links to actin cytoskeleton in 2 ways:
a. VERTICAL: Attaches to ankyrin complex which in turn attaches to band 3 (transmembrane bicarbonate transporter).
b. VERTICAL: Spectrin and actin both attach to band 4.1R complex, which in turn ataches to glycophorin C (another protein).
3. Membrane deformability - HORIZONTAL: Affects RBC shape and depends on cell's elasticity. Due to spectrin (possibly unfolding and refolding of specific spectrin repeats).
4. Cell viscosity - Function of intracellular hemoglobin concentration such that there is a maximum dissipation of the cytoplasm during cell deformation at a defined hemoglobin concentration.
Disorders of Cell Shape:
1. Hereditary spherocytosis (HS) - Generally autosomal dominant. Arises from defect in spectrin, ankyrin, band3, or band4.2 (also from VERTICAL causes aka spectin-ankyrin-band3 and spectrin-actin-band4.1, all of which stabilize the bilayer). Less stable RBC membranes means a decrease in membrane surface area such that the cells change shape from biconcave to spherical. So, less deformable and so they get trapped in splenic microvasculature. Removal of the spleen is largely curative. Symptoms:
a. Hemolytic anemia
b. Jaundice
c. Splenomegaly.
d. Gall stones
2. Hereditary Elliptocytosis - Due to horizontal (spectrin-spectrin interactions) and thus from spectrin, protein 4.1R, or glycophorin defects. Problems with cell membrane elasticity. Results in anemia, jaundice, and splenomegaly.
Cell-cell adhesion
When cells adhere directly to one another (as opposed to using the extracellular matrix as an intermediary). Generally done through cell-adhesion molecules (CAMs) which mediate two types of cell-cell adhesion: homotypic (between two cells of the same type) and heterotypic (between two different cell types). CAMs can interact with each other within a plasma membrane (cis) or perpendicularly with other cells (trans). On cytosolic side, CAMs are linked to linker proteins which then link to the cytoskeleton.
Intracellular Junction (4 types):
1. Anchoring junctions: Ensure structural integrity of tissues.
2. Occluding junctions: Ensure that molecular traffic is routed through cells instead of around them.
3. Channel-forming junctions: To enhance chemical, electrical and hence functional coupling of neighboring cells.
4. Signal-relaying junctions: To enhance cell-cell communication such as synaptic junction or cell-cell interaction through transmembrane ligand-receptor pairs (like Delta-notch).
CAMs: 4 types - Cadherins, Integrins, Immunoglobulin superfamily, Selectins
1. Cadherins are found in two types of anchoring junctions, adherens junctions and desmosomes. 3 types in adherens junctions:
a. E-cadherin - Epithelial
b. N-cadherin - Neural tissue
c. P-cadherin - Placental tissue
d. Structure of the above three "classic cadherins". N-terminal domain is extracellular and site for cell-cell adhesion which requires extracellular calcium. C-terminal domain is intracellular and interacts with cytoskeleton (mainly ACTIN) via alpha- or beta-catenin. Participates in both cis and trans interactions.
e. Classic Cadherins drive tissue differentiation via changes in number of E-cadherins which perpetuate a epithelial-mesenchymal transition where non-motile cells become motile. Important for NEURAL CREST migration!!!
f. Desmosomal cadherins (eg desmoglein and desmocollin). Differ from classical cadherins because bind to plakins instead of catenins and to intermediate filaments instead of actin.
1. Pemphigus vulgaris - Autoimmune skin blistering. Auto-immune bodies form against desmoglein which disrupts skin and mucous membrane integrity which creates spaces between cells which causes blistering. FLACCID blisters when compared to Bullous pemphigoid.
2. Bullous pemphigoid - Auto-immune directed at hemidesmosomes (in which integrins attach cells to the basal lamina). Causes skin blisters that are TENSE compared with Phemphigus vulgaris.
Cell-Matrix Adhesion
When cells adhere indirectly to each other using the extracellular matrix as an intermediary via adhesion receptors generally belonging to the INTEGRIN superfamily (eg fibronectin and laminins).
1. Structure - Heterodimers with 8 possible alpha-subunits and 18 possible beta-subunits.
2. Mechanism -
a. Connect to ACTIN via intermediate linker proteins (talin or vaniculin) attached to their C-terminal domains.
b. Active and inactive configuration. Generally binding with extracellular ligand results in change from inactive to active state (unfolds the legs) and subsequent binding to linker protein, but reverse is possible (inside-out activation).
c. Degree of adhesion depends on number of active integrins (not on strength of individual interactions).
d. Loss of this results in apoptosis.
1. Muscular Dystophy - Caused by defects in dystroglycan complex (DGC) which functions to connect actin cytoskeleton to sacrolemma (muscle cell plasma membrane). DCG binds to dystrophin in cytoplasm (which is bound to actin) and laminin in extracellular matrix.
a. Duchenne Muscular Dystrophy - Mutation in DMD gene (encodes dystophin) on X-chromosome. Starts in early childhood with profound weakness and muscle wasting with eventual involvement of all voluntary muscles and death due to respiratory arrest or congestive heart failure (survival into adulthood is uncommon).
b. Becker Muscular Dystophy - Same as Duchenne but milder and can survive to adulthood.
c. Can also be caused by post-translational problems of DGC.
d. Can also be associated with neuropsychiatric and behavior abnormalities, as well as cognitive dysfunction.
Actin cable growth
Actin exists in 2 forms, filamentous and globular. Globular actin contains a binding site for ATP. When it is bound by ATP (nucleation) it forms small collections of globular monomers called protofilaments. Several protofilaments bind together in alpha-helical filaments which have an inherent polarity (new actin monomers added to plus end while ATP binds to the negative end, which tends to be stable).
Arp2/3 complex - Attach the negative end of one actin cable to the middle of another, forming a branched, gel-like array.
Formins - Nucleate straight, unbranched filaments that can be cross-linked to form parallel bundles.
Amoeboid Migration
1. Extension - Actin polymerization pushes the membrane forward and away from cell body through generation of protruding structures (Filopodia = migrating growth cones and fibroblasts, Lamellipodia = epithelial cells, fibroblasts, neurons, Pseudopodia = amoebas and neutrophils). The rearrangement of the cytoskeleton is under control of Rho family monomeric GTPases (including Cdc42, Rac, and Rho). These have GDP bound and are inactive until plasma membrane bound guanine nucleotide exchange factors (GEFs) catalyze the exchange from GDP to GTP, activating them. Rho GTPases then associate with plasma membrane and activate Arp2/3 complex and Formins (among others).
2. Adhesion - Integrins connect the actin cytoskeleton to the non-moving extracellular matrix. The integrins also have some signalling properties.
3. Translocation - Force generated by myosin motor proteins that crosslink actin filaments contracts the actin cytoskeleton and moves the trailing edge of the cell (containing most of the cytoplasm) forward. This also weakens integrin interactions.
Degradation of ECM proteins
Mechanism -
1. Extracellular by Matrix Metalloproteases (require Ca++ or Zn++) or Serine proteases (have serine in active site).
2. Some proteins require transport into lysosomes for completion of degradation process.
Purpose - Generally, cell migration is promoted by degradation of ECM proteins for the following reasons:
1. Clearing a migratory path.
2. Exposing ends of ECM molecules for cell binding
3. Promoting cell detachment from ECM scaffold
4. Releasing signal peptides that stimulate migration.
Regulation -
1. Proteases are secreted in inactive form and need to be activated (eg plaminogen, a serine protease, needs to be cleaved/activated by tissue plasminogen activators (tPA)).
2. Proteases can be bound to cell surface receptors, limiting their actions to directly around a cell (eg urokinase-type plasminogen activators (uPA) are bound to growing tip of axons or other migrating cells).
3. Cell can secrete protease inhibitors (eg inhibitors of metalloproteases (TIMPs) and serine protease inhibitors called serpins).
Synthesis of mature collagen
1. Initially synthesized as pre-procollagen which has a signal peptide on N-terminus designating secretion. This localizes ribosome to RER where the peptide is extruded into the RER and the signal peptide is cleaved off, forming pro-alpha chains of collagen.
2. Some lysine and proline in the pro-alpha chains are hydroxylated to form hydroxylysine (via lysyl hydroxylase) and hydroxyproline (via prolyl hydroxylase). Both of these require ascorbic acid (Vitamin C), the deficiency of which can cause scurvy.
3. Glucose or glucose galactose dimer is added to some lysine residues.
4. Intra- and intermolecular disulfide bonds are formed between cysteine residues at the C-terminal of 3 alpha-chains via disulfide isomerase, creating a globular domain. This ensures the correct selection of alpha-chains, aligns the alpha-chains in the correct way for Gly-X-Y pattern, and prevents premature association of procollagen into fibrils.
5. The 3 chains are zipped up to form rod-like structure of procollagen and are secreted from the Golgi apparatus into ECM.
6. In ECM, proteases cleave the globular domain off procollagen, creating tropocollagen. Adjacent collagen molecules self-assemble into parallel, staggered arrangements.
7. Covalent cross linking between collagen fibrils is achieved by forming intermolecular lysine-lysine bridges via lysyl oxidase (requires copper as cofactor).
Osteogenesis Imperfecta
Generalized disorder of connective tissue caused by reduction in or abnormality of procollagen type I. Mutation in COL1A1 or COL1A2 cause different types. Manifested by bone fragility, blue sclerae, and other manifestations. Presents with h/o fracture with minimal trauma, familial short stature, perinatal death (OI type II), presenile hearing loss. Dentinogenesis Imperfecta (brittle teeth) present in 30% of cases. 4 subtypes (all are autosomal dominant unless otherwise specified):
1. OI type I - Mildest and most common. Bone fragility with fractures with mild trauma pre-puberty. Blue sclera or with purple/gray tint.
2. OI type II - Most severe often lethal in perinatal period (respiratory problems). Numerous fractures prenatally and during birth causing skeletal deformity. Sclera blue or with purple/gray tint.
3. OI type III - Bone fragility and severe deformity. Fractures at birth. Short limbs and stature. Sclera blue. Respiratory problems caused by rib deformity. Autsomal RECESSIVE.
4. OI type IV - Between I and III in severity. Bone fragility mostly before puberty. Short stature. Sclera WHITE. Mild to moderate bone deformity.
Ehlers-Danlos Syndrome (EDS)
1. EDS type I - Autosomal dominant characterized by hypermobile joints w/ associated fractures, fragile skin with easy bruisability, delayed wound healing, cigarette-paper scars, velvety skin, aortic root dilatiation, mitral valve prolapse, and intestinal diverticula. Mutation in TYPE V COLLAGEN.
2. EDS type II - Milder on spectrum but same disease as type I.
3. EDS type III (hypermobility type)- Patients have benign joint hypermobility and hyperextensible skin. Chronic pain with acute dislocations or osteoarthritis. Generally no life-threatening complications. Mutation in TYPE III COLLAGEN.
4. EDS type IV (vascular type) - Similar to type I but much more severe with recurrent arterial aneurysm, spontaneous intestinal rupture, uterine, bladder, and rectal prolapse, inguinal hernia, and pneumothorax. Neonates with clubfoot and congenital hip dislocation. Median age of death = 48. Mutation in TYPE III COLLAGEN.
5. EDS type VI (kyphoscoliotic form) - Similar to type I but also has scoliosis and ocular involvement (scleral fragility and increased risk of globe rupture). Intelligence normal and often normal life span with increased risk of rupture of arteries and respiratory compromise. Mutation in LYSYL OXIDASE gene. AUTOSOMAL RECESSIVE.
Notch signaling pathway
Mediates interaction of adjacent cells and thus accentuates differences between adjacent cells (for example lateral inhibition).
Structure of notch receptors: Single-pass with epidermal growth factor (EGF) like repeats in extracellular space.
Synthesis of notch receptors: Synthesized as one large peptide but cleaved such that they form a heterodimer.
Signaling: Ligands are called DSL ligants (Delta and Jagged).
1. When they bind notch receptors, two cleavages of the receptor releases an intra-cytoplasmic peptide (the notch tail).
2. The notch tail travels to the nucleus where it binds a transcription repressor called CSL (CBF1 in humans) thereby converting it into a transcription activator. The activated genes are also transcriptional regulators (note that in the absense of the notch tail, CSL binds co-repressors which silences notch target genes).
Functions: Retina, nervous system, hematopoiesis.
1. CADASIL syndrome - Cerebral arteriopathy, autosomal dominant, with subcortical infarcts and leukoencephalopathy.
2. Alagille Syndrome - Mutation in JAG1 or NOTCH2 (this has renal problem in addition to the below). Autosomal dominant. Symptoms:
a. Intrahepatic biliary hypoplasia leading to neonatal jaundice.
b. Cardiac abnormalities (pulmonic and peripheral valvular stenosis).
c. Skeletal abnormalities (Butterfly vertebrae and decrease in interpedicular distance in lumbar spine).
d. Ocular abnormalities (posterior embryotoxon, retinal pigment changes).
e. Characteristic facial features (broad forehad, pointed mandible, bulbous tip of nose).
Cilia Structure and Synthesis (structure and synthesis of basal bodies)
Structure: Microtubule based, have highly organized axoneme (think axon) which is extended from a modified centriole, known as the basal body, which anchors the axoneme in the cell. The axoneme is surrounded by a membrane continuous with plasma membrane but has different membrane receptors and ion channels.
1. Motile Cilia - 9+2, 9 outer doublet microtubules with a central pair. Rarely there is a 9+0 configuration.
2. Non-motile (sensory) Cilia - 9+0 configuration (except for kinocilium of hair cells which are 9+2).
Basal Body: Located at base of cilia and are platform on which cilia are constructed. Can be thought of as specialized cilia.
1. Structure - Cylindrical with 9 triplet microtubules blades arranged in the shape of a barrel. Each blade consists of a complete microtubule (A) with a second partial microtubule (B) grown off the first with a third partial microtubule (C) grown off the second.
2. Synthesis - 2 ways:
a. New centrioles are created during cell division (each cell has 2, duplicates them to make 4, then divides to get back to 2).
b. Lots of basal bodies can form at once from deuterosome or generative complexes.
3. Function -
a. Cilum orientation via basal foot which protrudes in a direction oriented to cilia beating. Also striated rootlet which protrudes towards cell interior.
b. Key for ciliogenesis and are required to act as template for axoneme (array of 9 microtubules).
i. Basal body moves to cell surface where it integrates with cell cortex and associates with a membrane bound vesicle.
ii. This complex fuses with plasma membrane and the basal body then acts as a last stop for proteins needed for cilial growth.
Intraflagellar Transport (IFT)
Method by which cilia and flagella transport axonemal precursors from their sites of synthesis in the cytoplasm to the ciliary tip or vice versa (as there is no way to synthesize anything in the cilia itself).
Antrograde transport (to the tip) - Along microtubules via kinesin 2 motor proteins.
Retrograde transport (from the tip) - Along microtubules via dynein 2.
Function of Motile Cilia (and how they work) + Disorders
Ciliary Movement -
1. Hydrolysis of ATP leads to formation and breaking of dynein cross bridges between A tubule of one doublet and the B tubule of the adjacent doublet.
2. As the microtubules slide on one side of the axoneme, the doublets on the other side move in the opposite direction, causing the cilium to bend.
Functions: Respiratory tract (mucus clearance), Ependymal cells lining ventricles of the brain (CSF movement), Female reproductive tract (move ovum).
1. Primary Cilia Dyskinesia (PCD, the immobile cilia syndrome) - Autosomal recessive with respiratory disress perinatally, chronic upper respiratory infection, and abnormal left-right asymmetry (situs inversus) in 50% of affected patients. Can have decreased fertility.
2. Kartagener's Syndrome - Triad of sinusitis, bronchiectasis, and situs inversus (may be same as PCD). This has led people to see that left-right axis determination is genetically related to ciliary functions. 2 theories:
a. Nodal Vesicular Parcel (NVP) - Vesicles full of Sonic Hedgehog and retinoic acid are secreted on the right side of Henson's node but are swept to the left side via movement of the motile cilia in the node. The vesicles break open and are sensed by cilia to the left of Hanson's node.
b. Two Cilia Model - Motile cilia at Hansen's node create a leftward ciliary flow, which is sensed by immotile cilia on the left side of the node (acting as mechanical sensors).
Function of Primary (nonmotile) Cilia + Disorders
Universal to every cell.
1. Sensory abnormalities -
a. Olfactory bulb - Odorants binds to receptors on cilia which open ion channels leading to APs and olfaction. Defects in cilia can cause anosmia.
b. Retina - Rod and cone photoreceptors have modified cilia that sense and transduce light signals. Also, consist of 2 parts, outer segment (with photopigment) and inner segment (with translational machinery) and a cilia connects the two and relies on intraflaggelar transport (IFT) to transport stuff between the two. Defects in IFT results in photoreceptor death.
2. Obesity -
a. Bardet-Biedl Syndromes (BBS) - Syndromes characterized by obesity, retinal rod-cone dystrophy, polydactyly, cognitive impairment, hypogonadism, urogenital malformation, and neuropathy. Caused by defects in IFT genes (obesity caused by disruption of neurons in hypothalamus.
3. Polycystic Kidney disease - Adult Polycystic Kidney Disease (APCKD) - Autosomal DOMINANT. Multiple cysts in kidneys, liver, and pancreas. Variable adult-onset kidney disease resulting in end-stage renal disease. Caused by defects in Polycystin-1 (PC1) or polycystin-2 (PC2). Explained below:
a. Normally, cilia extend off renal luminal epithelium and detect fluid flow through the nephron via PC1. Activation of PC1 leads to open PC2, a calcium channel, which goes onto activate a intracellular cascade.
b. When there is a defect in PC1 or PC2, the cells falsely detect that there is decreased fluid flow, resulting in an overgrowth of epithelial cells that manifests as cystic diseases of the kidney.
4. Pallister-Hall Syndrome (PHS) - Autosomal DOMINANT disorder characterized by hypothalamic hamartoma, bifid epiglottis or laryngeal cleft, pulmonary segmentation anomalies and polydactyly. Caused by mutation of GLI3 (the Gli 3 protein interacts with downstream signaling molecules of Sonic Hedgehog and is localized in the cilia tip via IFT).
a. Can also be caused by a lack of Shh signaling. There are two forms of Gli3, an activator and a repressor. When no SHH signal, repressor form dominates and (after retrograde IFT) represses downstream SHH pathway. When SHH signal is there, SMO travels to cilia tip via anterograde transport and causes more activator Gli3 which is then retrograde transported via IFT which activates downstream SHH proteins.
Hox code
Important for specifying location of limb development relative to anterior-posterior (AP) axis. Also have to do with forearm and hindarm identity.
Lateral plate mesoderm
Bone and connective tissues are derived from this. Fibroblast growth factors (FGF) are expressed here and help initiate limb bud outgrowth. The region at the apex of a limb bud, called apical ectodermal ridge (AER) expresses a different FGF.
Proximal-Distal Limb Growth
Apical ectodermal ridge (AER) is located at the apex of the limb bud. Early removal of the AER results in severe truncation while later removal only affects distal structures. Fgf2, 4, and 8 are expressed in the AER and maintain growth in the proximal-distal direction (removal of AER can be compensated by adding these 3 growth factors).
Progress zone - Located just proximal to the AER, it is a region of undifferentiated mesenchyme which specifies distal or proximal identity of bone. Cells that spend short time in progress zone form proximal bone, while spending a longer time means they will end up as distal bone.
Dorsal-ventral specification of limbs
Dorsal identity - Wnt-signaling pathway. Wnt7a induces Lmx1 expression.
Ventral identity - Default identity. Homeobox gene engrailed-1 (En-1).
Anterior-posterior specification
(Anterior is thumb, posterior is pinky).
Zone of polarizing activity (ZPA) - Confers posterior identity of limb. Secretes Sonic Hedgehog (Shh). Hox8 gene positions ZPA. Maintenance of Shh is via Fgf4 from Apical ectodermal ridge (AER) and the reverse is true (Fgf4 is maintained by Shh).
a. Early limb control region (ELCR) - Regulates the expression of the HoxD cluster in early embryonic development.
b. General Control Region (GCR) - Later in development, takes over from ELCR and decreases HoxD1 expression while upregulating Hoxd11, Hoxd12, and Hoxd13 which in turn simulate Shh production and secretion by ZPA.
Individual digits - Formed via apoptosis of interdigital mesenchyme.
Congenital Limb Malformations
1. Holt-Oram Syndrome - Caused by mutations in TBX5 gene. Characterized by upper limb malformations (most common are triphalangeal thumbs) and congenital heart disease (most commonly atrial septal defect).
2. Mutations in GLI3 (a member of Shh signaling pathway) cause brain and hand malformations such as Greig cephalopolysyndactyly syndrome (macrocephaly and postaxial polydactyly), the Pallister-Hall syndrome (hypothalamic hamartoma and polysyndactyly) and the acrocallosal syndrome (absense of corpus callosum, limb deformities, and other midline defects).
3. Mutations in LMX1B cause nail-patella syndrome, which is characterized by nail dysplasia, absent or hypoplastic patella, and nephropathy (makes sense because nails and patella are dorsal structures).
Brain development
Formed from rostral end of neural tube.
1. Anterior-posterior (rostral caudal) - Segmentation creates compartments called neuromeres. Defined by Hox genes.
a. Forebrain - Mediated by Homeobox gene Goosecoid and Otx1.
2. Ventral patterning - Due to Sonic Hedgehog (Shh) expressed in the prechordal mesendoderm and the notochord (which are adjacent to the prosencephalon and the rest of the brain, respectively).
3. Dorsal patterning - Via Bone morphogenic proteins (BMPs), which are part of TGF-beta gene family, and are produced in the non-neural ectoderm including the dorsal roof of the forebrain and the neuraxis. Also induce Msx1.
4. Combination of Shh and BMPs lead to differential expression of Dbx, Irx, and Nkx which specify neural differentiation into Dorsal, Ventral, and motor neurons.
5. Holoprosencephaly (HPE) - Can be caused by defect in Shh gene (among other things). Characterized by presence of a single cerebral vesicle instead of 2 distinct hemispheres. Associated with craniofacial abnormalities of the midline (cyclopia, cebocephaly/single ventricle, single proboscis, cleft lip and palate, hypotelorism, single central incisor).
Sonic Hedgehog (SHH) Signaling Pathway
1. A signal sequence at the amino terminal end of Shh is cleaved.
2. This triggers the cleavage of the remaining peptide into 2 halves.
3. A cholesterol is put on carboxyl-terminal residue representing the amino-terminal half of the original molecule. Palmitic acid is added to the carboxyl end of the molecule via an ester bond.
1. PATCHED1 (PTCH) is the cell surface receptor for Shh. Without Shh present, PTCH is bound to Smoothened (SMO).
2. When Shh binds PTCH, PTCH releases SMO.
Holoprosencephaly - Can be caused by defects in Shh, PTCH, SMO, deficiencies in cholesterol synthesis (Smith-Lemli-Opitz Syndrome), statins, Veratrum (inhibits SMO).
Gompertz Plot
Plots mortality rate vs. age logarithmically to get a linear plot (means mortality rate increases exponentially).
Equation: m(x) = Ae^(Gx) where m(x) is mortality rate at age x, A is initial mortality rate at age 0, and G the exponential rate of increase of mortality (aka Gompertz constant).
Mortality rate doubling time (MRDT): loge(2)/G (for human populations it is about 8).
Max life span = 15MRDT.
Between different populations: Variable in early life but converge later in life.
Wear and Tear Theory of Aging
Aging is an inevitable consequence of life itself, caused by accumulation of toxic metabolites, damage to various molecules in the body which results in deterioration of structure and function.
Wear and Repair (or Optimization) Theory of Aging
Patterns of senescence represent a balance between wear and repair. The wear and tear sustained through life is constantly being repaired at increasing energy costs. As life advances, the balance can no longer be sustained.
Mutation accumulation theory of aging
Over the course of evolution, many deleterious mutations have accumulated in the germ line that tended to decrease human reproductive fitness. Any novel mutation that can supress these mutations beyond reproductive years will increase the fitness of the species. The aging phenomena is a consequence of the cumulative effect of deleterious mutations and "suppressor" mutations.
Antagonistic Pleiotropy Theory of Aging
Specific alleles at many loci are selected over the course of evolution because they increase the reproductive fitness of the species. These same alleles may have a paradoxically deleterious effect later in life, but such effects are "selection-neutral" for the species.
Hutchinson-Gilford Progeria Syndrome (HGPS)
Characterized by progressive aging beginning at 12 months of age following an infancy marked by failure to thrive. Death generally from atherosclerosis and associated cardiac or cerebrovascular complications.
Caused by a mutation in progerin or LMNA which is a nuclear lamin called lamin A. The mutation is in a splice site that results in the loss of 150 nucleotides in the final mRNA.
Normal Lamin A -
a. Synthesized as Prelamin A which has CSIM (cysteine, serine, isoleucine, and methionine) on carboxyl end.
b. CSIM signals farnesyltransferase (FT) to attach a farnesyl group onto the cysteine.
c. At this, the SIM is cleaved off by ZMPSTE24 and the carboxyl terminal is capped with methyl group.
d. ZMPSTE24 cleaves the terminal 15 amino acids to form mature lamin A.
Abnormal Lamin A (in HGPS):
a. ZMPSTE24 cannot do the 2nd cleavage, leading the persistence of the farnesyl group and thus disorganization of the nuclear lamin.
b. The disorganization can be observed as nuclear blebbing and there is a disruption of gene regulation, epigenetic control, mRNA trafficking, protein function/turnover and DNA replication and repair.
Retinoblastoma (RB)
Classic example of 2-hit hypothesis. There are 2 mutations needed for the disease. Two types:
1. Familial Retinoblastoma (RB) - 40% of cases. Autosomal dominant. Age of onset is early infancy and there are usually multiple tumors and bilateral involvement. One mutation is inherited and so all somatic cells have it, while the 2nd one happens multiple times in different places. Each time the 2nd one happens, a tumor forms.
2. Sporatic RB - 60% of cases. Late onset (requires more time) and generally only one tumor. This is because all person's cells are initially normal and 2 mutations need to happen in the same cell for a tumor to form.
Loss of heterozygosity (LOH)
Method for detecting loss of a gene in a tumor.
1. Find a RFLP loci at which the patient is heterozygous.
2. A tumor suppressor gene show loss of an RFLP allele in the tumor when compared to normal tissue.
This is because there are 4 ways a somatic mutation can get rid of a tumor suppressor:
1. Local mutation events
2. Somatic recombination
3. Chromosomal loss followed by duplication
4. Chromosomal loss
In all but local events (I THINK, check questions?), the heterozygousity is lost.
Proto-oncogenes vs. tumor suppressor genes
do this you idiot
M-phase promoting factor (MPF)
There's a different pair for each part of the cell cycle (G1, S, G2, and M).
Structure: Composed of two components -
a. Cyclin - Regulation. Increases throughout interphase and maxes during mitosis (promoting M-phase) at which point it activates M-phase promoting factor (MPF). When it declines MPF activity falls off almost immediately.
b. Cyclin-dependent protein kinase (CDK) - Catalysis.
Regulation Cycle:
a. Initially (just after M) no cyclin is present, so CDK is inactive.
b. Cyclin starts to be synthesized and forms complexes with CDK. These complexes are still inactive because they are phosphorylated on Tyr15.
c. Phosphorylation of Tyr100 and dephosphorylation of Tyr15 activates the cyclin-CDK complex.
d. Active CDK phosphorylates phosphatase (activating more CDK-cyclin complex)
e. Active CDK phosphorylates destruction box recognition protein (DBRP), a subunit of anaphase-promoting complex (this adds ubiquitin molecules to cyclin via ubiquitin ligase).
f. Ubiquitination of cyclin causes it to be degraded by proteosomes, leaving CDK inactive.
Other regulation:
a. Cyclin-dependent kinase inhibitors (CKIs) - Inhibit the activation of G1 and S phase cyclin/CDK complexes by regulating the synthesis of their subunits.
b. G1-cyclin/CDK complexes phosphorylate CKIs to target them for degradation.
Function: Causes progression through M-phase (through phosphorylation).
a. Nuclear lamins - Depolymerizes nuclear lamina leading to breakdown of nuclear membrane.
b. Condensin complex - Increases DNA coiling activity of condensin subunits responsible for chromatin condensation.
c. Myosin light chain - Prevents myosin from interacting with and sliding along actin filaments (protects against premature cytokinesis).
d. Lots of others.
e. S-phase CDK complexes activate existing pre-replication complexes while preventing formation of new pre-replication complexes.
Retinoblastoma Protein (RB)
Negative regulator of the cell cycle.
1. When active (unphosphorylated), Retinoblastoma Protein (RB) inactivates E2F protein (needed to express genes required for DNA replication).
2. G1-cyclin/CDK phosphorylates (and thus inactivates) RB resulting in activation of E2F and thus increased expression of CDK2 and S-phase cyclins.
RAS/MYC pathway
Gets through G1 checkpoint by activating E2F.
1. Growth factors or other mitogens bind to a receptor.
2. RAS is activated.
3. RAS activates Raf serine/threonine kinase, which in turn activates MAP kinase, which goes on to activate transcription factors.
4. Transcription factors increase transcription of target genes, for example the proto-oncogene MYC.
5. MYC increases cyclin D, SCF subunit, and E2F gene.
6. Cyclin D leads to increase of G1-CDK activation -> retinoblastoma protein inactivation -> increased E2F activity -> S phase.
7. SCF subunit increases p27 degradation, increasing G1/S-CDK activation -> retinoblastoma protein inactivation -> increased E2F activity -> S phase.
1. Mitogen-independent activation of RAS leads to easy E2F and thus S-phase.
2. Decreased activity of RB or p16 is a problem (both are tumor suppressors).
3. Increased activity of cyclin D1 or CDK4 genes (both are proto-oncogenes).
Positive and negative regulators of cells death (genetics) conserved from Nematodes
Positive regulators of cell death: Ced-3 and Ced-4 (in nematodes) which map to CASPASES and APAF-1 respectively in humans. Ced-4 activates Ced-3. If either of these have inactivating mutations, then there is no cell death.
Negative regulator of cell death: Ced-9 (in nematodes) which is BLC-2 in humans. Activating mutations prevent cell death. Ced-9 represses ced-4.
If Ced-3 and Ced-9 are inactive, then no extra or less cell death, which suggests that most cells are actively expressing the ced-3 pathway of cell death but it is repressed by ced-9.
Molecular Apoptosis
Once a cell death signal, the transduction of that signal is modulated via BCL-2 family proteins. Bcl-2 and Bcl-X1 are anti-apoptotic/pro-survival while Bax, Bim, and Bad are pro-apoptotic. The relative amounts of these two classes of Bcl-2 family proteins determines whether the signal is transduced. Method of actions for Bcl-2:
1. Bcl-2 is localized to the mitochondrial membrane. To start apoptosis, generally the mitochondrial membrane potential is lost, ions are released, and larger molecules including cytochrome c are released (which directly leads to apoptosis). Bcl-2 inhibits the release of cytochrome-2.
2. Bax is thought to facilitate cytochrome-2 release.
Execution phase of apoptosis (Caspases):
1. Caspases are normally present in cells as inactive zymogens. Proteolytic cleavage of inactive procaspase results in a large and small subunit that combine to form a heterodimer which combines with another heterodimer to form an active caspase.
a. Proteolytic cleavage of caspases is catalyzed by other caspases.
b. Enough procaspases together can be fully activated by themselves.
2. Initiator caspases are activated first. These contain death effector domains (DED) or caspase recruitment domains (CARD).
3. Initiator caspases cleave (and activate) effector caspases (which contain short prodomains).
4. Active effector caspases cleave all sorts of stuff, including inactivating inhibitors of apoptosis, disassembling cell structures, and deregulating the activity of various transduction proteins and homeostatic proteins.
Extrinsic Cell Death Pathway
Death receptors like Fas bind its ligand and then recruit, aggregate, and autocatalytically activate initiator procapases (involves interaction between death domains and death effector domains). Important for:
1. Deletion of activated mature T cells in periphery at the completion of an immune response.
2. Killing of target cells by cytotoxic T-cells and natural killer cells.
3. Killing inflammatory cells at "immune-privileged sites" such as the eye.
4. An increased concentration of soluble Fas (which may inhibit FasL-Fas reactions) a mutation in FasL occur in some patients with systemic lupus erthematosus.
5. Children carrying heterozygous mutation in Fas gene have autoimmune lymphoproliferative syndromes (ALPS).
Intrinsic Cell Death Pathway
1. Death stimuli cause Bax to accumulate on the mitochondrial outer membrane.
2. Bax with BH3-only proteins trigger release of cytochrome c from the mitochondrial inter-membrane space.
3. Cytochrome c forms a complex with Apaf-1 and procaspase-9 in the cytosol called an apoptosome.
4. The apoptosome results in the aggregation and autocatalytic activation of caspase-9.
5. Active caspase-9 initiates a cascade that results in the activation of effector caspases 3 and 7.
A tumor suppressor gene that encodes a transcription factor that acts a checkpoint to sense DNA damage.
1. If the DNA is repairable, p53 induces the expression of genes that produce cell cycle arrest.
2. If the DNA damage is too severe and cannot be repaired, p53 activates a cell death pathway.
a. p53 induces transcription of Bax (a pro-apoptotic Bcl-2 related protein that increases cytochrome c release from mitochondria).
Problems: If you don't have active p53, cells are less susceptible to death due to irradiation or DNA-damaging cells, thus allowing these cells to accumulate cancer-causing mutations (and thus obviously increasing rates of cancer).
Ataxia Telangiectasia
Caused by homozygous mutations in the ATM gene, which codes for serine protein kinase. ATM has two major roles in relation to DNA damage repair and apoptosis:
1. It is part of complex called BASC (BRCA1-associated genome surveillance complex) which includes other tumor suppressor proteins such as NBS, BRCA1, and BRCA2. BASC plays a role in double-strand break repair.
2. ATM phosphorylates TP53 at the Ser15 residue, activating its apoptotic function. It also indirectly phosphorylates the Ser20 of TP53 and thus prevents TP53 from being degraded.
3. So, a homozygous defect in ATM results in an abnormal cellular response to radiation-induced double-strand DNA break. Some mutations act as DOMINANT NEGATIVE.
Symptoms (onset in first decade of life):
1. Progressive telangiectasia (mainly on conj, ears, and face).
2. Cerebellar ataxia
3. Nystagmus, myoclonic jerks, dystonia, and athetosis
4. Recurrent pulmonary infection
5. Increased risk of lymphoreticular malignancy
Lab Abnormalities include:
1. Chromosomal translocation of 7 and 14
2. Low IgA and IgG2 due to thymic hypoplasia.
3. Elevated alpha-fetoprotein and carcinoembryonic antigen
4. Increased radiosensitivity
5. Pathology showing loss of purkinje, granule, and basket cells in cerebellar cortex and neurons in the deep cerebellar nuclei, loss of anterior horn cells in spinal cord and of DRG cells associated with posterior column spinal cord demyelination.
Type of cancer commonly mutated in: Breast, ovarian, other carcinomas
Cancer predisposition syndrome: None
Type of molecular function: Receptor tyrosine kinase
Molecular pathways it is found in: Epidermal growth factor signaling
Effect on cell growth: Proto-oncogene
Importance: Amplification and/or overexpression of HER-2 confers poor prognosis to breast cancer and antibodies directed against it (trastuzumab or Herceptin) have been shown to be effective in some patients with HER-2 amplification/overexpression.
Type of cancer commonly mutated in: Medullary thyroid carcinoma
Cancer predisposition syndrome: Multiple endocrine neoplasia type II.
Type of molecular function: Receptor tyrosine kinase
Molecular pathways it is found in: Glial-derived neurotrophic growth factor.
Effect on cell growth: Proto-oncogene
Importance: While gain of function mutation (germline or somatic) increases the risk of oncogenesis, loss of function results in Hirschsprung disease.
Type of cancer commonly mutated in: Lymphhomas (c-myc), neuroblastoma (n-myc), and small cell lung carcinoma (l-myc).
Cancer predisposition syndrome: None
Type of molecular function: Transcriptional regulators
Molecular pathways it is found in: Common to many pathways
Effect on cell growth: Proto-oncogene
Importance: c-myc activation by means of chromosomal translocations are well-documented in Burkitt's lymphoma; N-myc gene amplification in neuroblastoma is associated with poor prognosis; L-myc gene amplification is found in many small cell lung carcinomas. Sometimes, amplification of myc can take cytogenetic form of double minute chromatin bodies or as homogeneous staining regions (HSR).
Type of cancer commonly mutated in: B-cell lymphoma
Cancer predisposition syndrome: None
Type of molecular function: Anti-apoptosis protein
Molecular pathways it is found in: Apoptosis pathway
Effect on cell growth: Proto-oncogene
Importance: Frequently activated in B-cell lymphoma due to a chromosomal translocation between 14 and 18.
Type of cancer commonly mutated in: Many, particularly pancreatic and colorectal (k-ras), myeloid leukemia (n-ras) and bladder cancer (h-ras)
Cancer predisposition syndrome: None
Type of molecular function: Signal regulator (p21 GTPase)
Molecular pathways it is found in: Many, especially tyrosine kinase
Effect on cell growth: Proto-oncogene
Importance: Some mutations may be associated with poorer prognosis.
Type of cancer commonly mutated in: Chronic myelogenous leukemia
Cancer predisposition syndrome: None
Type of molecular function: Non-receptor tyrosine kinase
Molecular pathways it is found in: Non-receptor tyrosine kinase pathway
Effect on cell growth: Proto-oncogene
Importance: Part of a fusion protein, Bcr-Abl, created as a result of the chromosomal translocation between chromosomes 9 and 22; the increased nuclear tyrosine kinase activity is necessary and sufficient for the causation of chronic myelogenous leukemia; the highly effective and selective tyrosine kinase inhibitor Imatinab or Gleevac is a Bcr-Abl antagonist that represents the first targeted therapy for neoplasia.
Type of cancer commonly mutated in: Retinoblastoma, osteosarcoma
Cancer predisposition syndrome: Familial retinoblastoma
Type of molecular function: Cell cycle regulator
Molecular pathways it is found in: Cell cycle progression
Effect on cell growth: Tumor suppressor gene (TSG)
Importance: Un-phosphorylated-pRB binding to E2F represses the expression of genes needed for G1-S progression of the cell cycle; DNA tumor viruses cause cancer in animals by inactivating RB.
TP53 (P53)
Type of cancer commonly mutated in: Wide spectrum
Cancer predisposition syndrome: Li-Fraumeni Syndrome
Type of molecular function: Transcriptional regulator; pro-apoptotic protein
Molecular pathways it is found in: Cell cycle progression; apoptosis pathway; metastasis
Effect on cell growth: Tumor suppressor gene (TSG)
Importance: Perhaps most commonly mutated gene of all cancers; constitutional deletion causes Li-Fraumeni sydrome (autosomal dominant which confers high risk for breast and brain cancer, soft tissue sarcoma, osteosarcoma, adenocortical carcinoma, and leukemia), p53 is the target used in many experimental gene therapy protocols.
Type of cancer commonly mutated in: Colorectal cancer
Cancer predisposition syndrome: Familial polyposis (or adenomatosis polyposis coli)
Type of molecular function: Signal regulator, mitotic spindle binding protein
Molecular pathways it is found in: Wnt signaling; cell cycle (mitotic spindle formation)
Effect on cell growth: Tumor suppressor gene (TSG)
Importance: Mutated in 80% of all colorectal cancers; constitutional mutation results in various familial polyposis syndromes which account for 1% of all colorectal cancers.
Details on the Wnt pathway and APC's role:
1. Wnt proteins are signaling glycoproteins which bind to Frizzled family of cell surface receptors.
2. When Wnt proteins bind Frizzled, Frizzled associated with LDL receptor-related proteins (LRP) and the complex begins an activation pathway (there are multiple but we are only doing one aka the Canonical Wnt pathway).
3. Beta-catenin molecules are usually associated with E-cadherin.
4. Unassociated beta-catenin are degraded by a protein complex consisting of constitutively active enzyme GSK-3beta (glycogen synthase kinase 3beta), APC protein (adenomatous polyposis coli), scaffold protein axin which holds the complex together. The beta-catenin is bound and phosphorylated by the complex and then ubiqutinated which marks it for proteosomal degradation.
5. A wnt bound frizzled-LRP complex activates the protein Dishevelled.
6. Dishevelled inhibits the activity of GSK-3beta leaving cytoplasmic Beta-catenin unphosphorylated (and thus no ubiquitination or degradation).
7. Beta-catenin accumulated in the cytoplasm and eventually makes its way to the nucleus, where it activates target genes by displacing a co-repressor called Groucho and facilitating the regulatory protein LEF-1/TCF by serving as a co-activator.
8. One of the target genes for Beta-catenin is c-myc (a proto-oncogene).
Type of cancer commonly mutated in: Neurofibrosarcoma, brain tumors
Cancer predisposition syndrome: Neurofibromatosis type I
Type of molecular function: Signal regulator (GTPase-activating protein).
Molecular pathways it is found in: Many signaling pathways, especially receptor tyrosine kinase pathways.
Effect on cell growth: Tumor suppressor gene (TSG)
Importance: Constitutional mutation causes the autosomal DOMINANT disorder neurofibromatosis type I.
Type of cancer commonly mutated in: Acoustic neuroma, meningioma, glioma, schwannoma
Cancer predisposition syndrome: Neurofibromatosis type 2
Type of molecular function: Cytoskeletal protein
Molecular pathways it is found in: Cell adhesion
Effect on cell growth: Tumor suppressor gene (TSG)
Importance: Mutation in the gene result in loss of coordination between growth factor signaling and cell adhesion
CDKN2A (P16ink4a)
Type of cancer commonly mutated in: Melanoma, glioma, leukemia, bladder cancer, head and neck cancer
Cancer predisposition syndrome: Familial melanoma
Type of molecular function: Cell cycle inhibitor
Molecular pathways it is found in: Cell cycle progression (cyclin-dependent kinase inhibitor)
Effect on cell growth: Tumor suppressor gene (TSG)
Importance: Constitutional mutations results in familial melanoma
Type of cancer commonly mutated in: Wilms Tumor
Cancer predisposition syndrome: Familial Wilms tumor (rare), WAGR syndrome, Denys-Drash syndrome
Type of molecular function: Transcription regulator
Molecular pathways it is found in: Transcription suppression of growth-inducing genes
Effect on cell growth: Tumor suppressor gene (TSG)
Importance: Deleted as part of the contiguous gene syndrome of WAGR (Wilms tumor, aniridia, genital hypoplasia, and retardation of growth and development); germline missense mutation results in Denys-Drash syndrome, which consists of Wilms tumor, ambiguous genitals, and nephropathy.
Type of cancer commonly mutated in: Basal cell carcinoma, medulloblastoma
Cancer predisposition syndrome: Basal cell nevus syndrome (Gorlin syndrome)
Type of molecular function: Cell surface receptor
Molecular pathways it is found in: Shh signaling pathway
Effect on cell growth: Tumor suppressor gene (TSG)
Importance: Constitutional mutation results in the Gorlin syndrome, which consists of basal cell nevus (and carcinoma), odontoid cysts and palmar and plantar cysts.
Type of cancer commonly mutated in: Renal cancer, pheochromocytoma
Cancer predisposition syndrome: Von Hippel-Lindau syndrome
Type of molecular function: Upiquitin ligase complex
Molecular pathways it is found in: Angiogenesis (prevents accumulation of hypoxia inducible factor-1, HIF-1)
Effect on cell growth: Tumor suppressor gene (TSG)
Importance: Constitutional mutation results in the Von-Hippel-Lindau syndrome, an autosomal dominant syndrome consisting of cerebral hemangioblastoma, retinal angioma, and renal cysts and carcinoma.
CDH1 (E-Cadherin)
Type of cancer commonly mutated in: Gastic cancer
Cancer predisposition syndrome: Familial gastric cancer
Type of molecular function: Cell adhesion molecule
Molecular pathways it is found in: CEll adhesion (an invasion suppressor)
Effect on cell growth: Tumor suppressor gene (TSG)
Importance: Constitutional mutation causes familial gastric cancer; loss of CDH1 function occurs late in many other cancers, and is likely to account for local invasion and metastasis in many late stage cancers.
Type of cancer commonly mutated in: Breast, ovarian, prostate, pancreatic cancer
Cancer predisposition syndrome: Familial breast and ovarian cancer
Type of molecular function: Mediator of double strand break repair, transcriptional regulator
Molecular pathways it is found in: DNA repair, transcriptional regulation
Effect on cell growth: Tumor suppressor gene (TSG) (DNA repair)
Importance: Germline mutation of BRCA1 results in familial breast and ovarian cancer
Type of cancer commonly mutated in: Breast, ovarian, prostate, pancreatic cancer
Cancer predisposition syndrome: Familial breast and ovarian cancer
Type of molecular function: Mediator of double strand break repair, transcriptional regulator
Molecular pathways it is found in: DNA repair, transcriptional regulation
Effect on cell growth: Tumor suppressor gene (TSG) (DNA repair)
Importance: Germline mutation of BRCA2 results in familial breast and ovarian cancer; heterozygous mutation BRCA2 causes Fanconi pancytopenia syndrome (consists of pancytopenia, radial anomaly, short stature, and increased risk of leukemia).
Type of cancer commonly mutated in: Lymphoma
Cancer predisposition syndrome: Ataxia telangiectasia
Type of molecular function: Protein kinase; mediator of double strand break repair
Molecular pathways it is found in: DNA repair; cell cycle progression
Effect on cell growth: Tumor suppressor gene (TSG) (DNA repair)
Importance: ATM phosphorylates p53 and BRCA1, thus mediating cell cycle control and double strand break repair, respectively; homozygous mutation results in the syndrome ataxia telangiectasia.
Type of cancer commonly mutated in: Colorectal and other gastrointestinal cancers, endometrial cancer, ovarian cancer, biliary cancer
Cancer predisposition syndrome: HNPCC (lynch syndrome)
Type of molecular function: Mismatch repair
Molecular pathways it is found in: DNA repair
Effect on cell growth: Tumor suppressor gene (TSG) (DNA repair)
Importance: HNPCC, also found in sporadic colorectal cancers.
One common arrangement of mass spectrometer (used to detect the protein contents of cells). Matrix-assisted laser desorption (MALDI), coupled with mass analyzed by time of flight (TOF).
- Biological molecules dispersed in a biological matrix.
- UV laser pulse ablated the matrix, carrying some molecules into the gas phase in an ionized form
- Molecules sent to a mass spectrometer
TOF Analyzer:
- Measures the time it takes ions of different masses to move from ion source to the detercotr.
- Requires that the starting time (time at which ions leave the ion source) is well-defined.
- Thus, ions are formed by pulsed ionization method (usually MALDI), or various kinds of rapid electric field switching are used as a 'gate' to release the ions from the ion source in a very short time.
Sensitivity of reflector:
- Mass resolution - several ppm for mass
- Sensitivity 1fmol (10-15 mol)
One common arrangement of mass spectrometer (used to detect the protein contents of cells). Liquid chromatography, electrospray ionization, ion trap.
Electrospray ionization (ESI): A solution is sprayed into the source chamber to form droplets. The droplets carry charge when they exit the capillary and as the solvent vaporizes the droplets disappear leaving highly charged analyte molecules.
Ion Trap Analyzer:
- Advantage of ms/ms (tandem mass spectrometry) gives sequence of peptide.
- Often combined with electrospray and liquid chromatography.
- Operates by storing ions in the trap which are then used for analysis
- A portion of ions are used to produce a mass spectrum
- For ms/ms experiments, one or more mass/charge species is individually fragmented for analysis
- Very robust and sensitive and relatively inexpensive. Most common method.
Mass spectrometry (uses)
1. Identification of individual purified proteins
2. Identification of modification state (e.g. phosphorylation)
3. Annotation of large proteomes
4. Identification of proteins co-purifying in complexes
a. Generate strain with an ORF fused to a purification (TAP) tag
b. Purify ORF-fusion protein using affinity tag
c. Identify complexes by mass spectrometry
5. Comparison of populations of proteins. Example is comparing the amount of proteins in normal vs. transformed cell lines.
a. Within a single population, you can't say that X protein is more abundant than Y protein because they ionize differently.
b. However, if you look at the same protein in different populations that are prepared the same way, you can determine relative abundance. The problem is that you need to separate the peaks so you can tell the difference between protein X in cell A vs protein X in cell B. This can be accomplished by using a different isotope in the different cell populations so that the proteins have different masses. There are 3 ways to do this:
i. Proteins are labeled metabolically by culturing cells in isotopically enriched (e.g. with 15N or 13C-labeled amino acids) or isotopically depleted media
ii. Proteins are labeled at specific sites with isotopically encoded reagents. The reagents can also contain affinity tags, allowing for the selective isolation of the labeled peptides after protein digestion.
c. Proteins are isotopically tagged by enzyme-catalyzed incorporation of 18O from H218O during proteolysis (by trypsin). Each peptide is thus mass-labeled at the C-terminal.
Protein microarrays
What I think is going on: you affix a bunch of something to the chip and wash it in cells that have been lysed and see what binds.
1. Generate a genomic collection of strains each expressing a different ORF as an ORF-fusion protein.
2. Purify proteins in cells in parallel
3. Fix proteins to a glass slide or other solid support
4. Assay for binding
1. Allows high throughput assays
2. Identification of multiple proteins that bind target in one experiment
3. Ideal for identifying binding targets of drugs
1. Not good for enzymatic activity
Activity-based protein profiling
Chemical probes that react covalently with enzymes of specific classes.
The ideal probe is:
1. Specific for a class of activity
2. General for all activities of that class
3. Reacts in a manner that correlates with catalytic activity
4. Displays minimal cross-reactivity with other protein classes
5. Possesses a tag for rapid detection and/or isolation
Advantages to combination therapy for cancer (different classes of chemotherapeutic agents)
1. Maximal cell kill within the range of toxicity tolerated by host.
2. Each component can target a different vulnerability of the cancer cells
3. Circumvent drug resistance that cancer cells can develop to individual chemotherapeutic agents.
Growth fraction
For cancer. The number of cells that are actively growing (being in G1, S, M, or G2, NOT in G0). On average, this is 20%, which implies that although tumor growth is exponential, the rate of growth (proportional increase of cancer cells over a fixed period) is constant. THIS IS ONLY TRUE FOR LIQUID CANCERS (like leukemia).
Rate of growth for "liquid" vs "solid" cancers
Liquid - The number of cells that are actively growing (being in G1, S, M, or G2, NOT in G0) aka growth fraction is, on average, 20%, which implies that although tumor growth is exponential, the rate of growth (proportional increase of cancer cells over a fixed period) is constant.
Solid - Initial growth tends to be exponential (with constant growth rate) but tends to decrease as the tumor gets bigger (so smaller tumors grow faster than larger tumors). Called Gompertzian Growth model. This implies that you need to eliminate all cancer cells to make a difference in 5-year survivals, because small tumors will get back up to size quickly. Also implies that you should focus on dealing with cancer cells early when a larger percentage of them are in cell cycle and thus vulnerable to chemotherapy.
Log Cell Kill Method
Systemic treatment scheme for cancer. Assumes that treatment will kill a fixed percentage of cells, not an absolute number. So, if we do 3 successive therapies that have a 90% kill rate, we will end up with a 99.9% kill rate of the initial cell population. Thus, treatments are given in cycles iterative over time. This also has the advantage of allowing non-cancer cells to recover from drug toxicity between cycles.
Biochemical changes in cancer cells (Warburg effect)
Warburg effect - The observation that tumor cells have a high rate of glucose utilization compared to normal tissues. Also, most of the glucose is converted to lactate suggesting submaximal activity of the Krebs cycle and oxidative phosphorylation, despite the presence of oxygen. Called anaerobic glycolysis.
So, generally, 4 characteristics in cancer cells:
1. Increased glucose intake
2. Increased lactate production
3. Increased glutamine uptake
4. Increased production of lipids and nucleic acids.
Three theories as to why the anaerobic glycolysis predominates:
1. In rapidly growing tumors, areas of hypoxia can develop due to failure of angiogenesis to keep pace. When this happens cells may be forced to go into anaerobic glycoloysis as an adaptation. A more permanent metabolic transformation that grants cancer cells independence from fluctuating oxygen supply can remove yet another obstacle for continued uncontrolled cell growth (a proto-oncogene).
2. Tumor cells maintain robust glycolysis to keep pools of glycolytic intermediate available for biosynthesis. The high lactate levels are just a byproduct of the high glycolytic rate. While other nutrients such as non-essential amino acids, fatty acids, and cholesterols are also available from extracellular fluid, their constant supply cannot be guaranteed and the ability for de novo synthesis needs to be assured with a constantly high glycolytic flux. This reflects the different goals of cancer cells, as long as there's enough glucose available, the cancer doesn't care about being efficient, it just cares about having biosynthetic intermediates.
3. Lactate may not be just a passive by-product of glycolysis but an "intended" product because its secretion into the extracellular space may cause surrounding normal cells to die while promoting degradation of the extracellular matrix. Both of these can enhance the likelihood for tumor invasion and metastasis. Suppressing LDH activity would impair tumorgenesis which has indeed been shown for some cancers.
Other biochemical quirks in cancer cells:
1. High glutamine levels needed - Cancer cells have high lipid synthesis needs. Thus, citrate is generally shipped out of the mitochondria to provide Acetyl CoA for cholesterol and fatty acid synthesis. This means that the Krebs cycle is cut short and cannot progress. Glutamine can be turned into alpha-ketoglutarate which is incorporated into the Krebs cycle after citrate, bypassing the problem entirely.
2. Lots of purine and pyrimidine synthesis in cancer cells, which require glycolytic intermediates to synthesize key amino acids like glycine and in generation of ribose-5-phosphate from pentose phosphate pathway.
How are these metabiolic changes mediated?
1. Continual activation of the phosphatidyl ionositol-3-kinase (PI3K) signaling pathway. PI3K results in the phosphorylation of the membrane phosphatidyl inositol (PI) to form PI phosphate (PIP), which promotes growth through activation of the Akt/mTOR pathway. Upregulation of PI3K is accomplished via Her-2/neu epidermal growth factor receptor (EGFR) and platelet-derived growth factor receptor (PDGFR).
2. HIF-1 increases angiogenesis and increases expression of glucose transporters and all glycolytic enzymes.
3. c-myc upregulates NUCLEOTIDE SYNTHESIS via IMP dehydrogenase, serine hydroxymethyl transferase, adenosine kinase, adenylate kinase-2, and PRPP-aminotransferase.
Types of Chemotherapy (and other drugs that attack cancer)
1. M-phase (microtubules inhibitors) - Vinblastine, vincristine, taxanes.
2. G1 phase - Glucocorticoids (prednisone)
3. S-phase - Antimetabolites and folate pathway inhibitors
a. 5-Fluorouracil (prodrug for an inhibitor of thymidylate synthase).
b. 6-Mercaptopurine (prodrug for inhibitor for purine synthesis)
c. Hydroxyurea (inhibitor of ribonucleotide reductase)
d. Thioguanine (guanine analogue)
e. Cytarabine, 5-azacytidine, gemcitabine (cytidine analogues).
4. Transition from S to G2 - Topoisomerase inhibitors (camptothecin, topotecan, irinotecan, Anthracyclines such as doxorubicin and daunorubicin, Epopodophyllotoxins such as etoposide).
5. G2 phase - Antitumor antibiotics (Dactinomycin)
6. Cell cycle nonspecific - Alkylating agents and platinum complexes.
a. Cyclophosphamide, mephalan, chlorambucil, thiotepa (guanine alkylators)
b. Nitrosoureas
c. Dacarbazine and procarbazine
d. Cisplatin and carboplatin
e. Bleomycin (double strand breaker)
Non chemotherapy:
1. Amino acid degraders - Asparaginase denies cancer cells access to asparagine.
2. Inducer of differentation - All-trans-retinoic-acid (ATRA) causes cancer cells to become better differentiated, slowing growth.
Immunotherapy (for cancer)
Tumor cells often express unique surface antigens. This means that if you can design monoclonal antibodies that attach to these antigens, you can induce an immune response to the cancer cells. Rituximab is a monoclonal antibody that is directed against the CD20 antigen found on mature B cells that is used to treat B-cell non-Hodgkins lymphoma. Conjugation of radioactive isotopes to anti-CD20 antibodies, such as iodine 131 tositumomab or yttrium-90 ibritumomab tiuxetan allows them to be used in radioimmunotherapy of similar tumors.
Growth Factor Receptor and Signal Transduction Antagonists (treatment for cancer)
Antibodies directed against growth factor receptors.
1. Cetuximab, which is directed against EGFR, and trastuzumab, which is directed against HER-2, have been shown effective against tumor expressing those receptors (colon cancer and breast cancer respectively).
2. Tyrosine kinase inhibitors such as gelfitinib, which inhibits EGFR-mediated signaling, and imatinib, which is directed against BCR-ABL fusion protein, are effective against small cell lung cancer, breast cancer, and chronic myelogenous leukemia.
Proteosome Inhibitors
Proteosomes can act as a safety valve for cancer cells because cancer cells build up large amounts of misfolded or abnormal proteins that, if allowed to accumulate, can result in cell death.
Bortezomib, a proteosomal inhibitor, has been used successfully in treatment of multiple myeloma.
Angiogenesis Inhibitors
1. Bevacizumab - A monoclonal antibody against VEGF-A successful vs. colon cancer and non-small cell lung carcinoma.
2. Thalidomide - An inhibitor or bFGF-induced angiogenic. Treats multiple myelomas.
DNA repair inhibitors
Based on synthetic lethality, the observation that many cancers get worse with some impairment of DNA repair but further inhibition of DNA repair leads to cell death.
Reason: Normally there are 6 DNA repair pathways. If one is gone, then there is an increased reliance on a 2nd. If that one goes too, then there's too much DNA damage and we get cell death.
Iniparib - Inhibitor of poly-ADP-ribose-polymerase-1 (PARP-1) (which is required for base-excision repair). Being tested for breast cancer. Since most breast cancer has defective double-strand break repair capabilities it is very dependent on other DNA repair mechanisms. By inhibiting base-excision repair, the actively dividing cancer cells get too much DNA damage and die.
Taste (Gustation)
Five tastes, salty (mineral content), sour (acidity), sweet (quick calories), bitter (poison), and umami (amino acids and glutamate). Innervated via glossopharyngeal nerve (only sweet and bitter) and facial nerve (all 5 sensations), as well as vagus.
Ion Channel Taste Receptors -
1. Salty taste receptor: An epithelial sodium channel (ENaC). When exposed to salt, influx of sodium causes depolarization. In the sodium channels, expression of ENaC is stimulated by aldosterone.
2. Sour taste: Less well known but probably several channels. H+ ion can pass through same ENaC for salty taste sensation and can interfere with the potassium channel. Also, another channel called transient receptor potential cation channel (TRPP2) is involved.
G-Protein-Coupled Taste Receptors -
1. Sweet Taste Receptor - T1R (taste 1 receptor) family of GPCRs. Two isoforms T1R2 and T1R3 interact to form heterodimers to bind to a tastant. Upon binding to sugar, the Gs-controlled pathway is activated resulting in increased cAMP (via activation of adenylate cyclase) which results in inhibition of potassium channels, gating (keeping open) a non-specific cation channel called cyclic nucleotide-gated (CNG) channel in the taste receptor cells. Both of these depolarize the cell.
a. Artificial sweeteners: Depolarize taste receptor cells via Gq phospholipase Cbeta pathway. Some protein called gustducin and an ion channel called TRPM5 interact to depolarize the taste cell.
b. Leptin, an adipokinetic hormone, lowers the sensitivity of sweet taste receptor cells (help with weight loss via decrease of "sweet tooth").
2. Umami Taste - Similar to sweet taste receptor but the heterodimer is T1R1 and T1R3 (not 2). Specific for L-glutamate. Signal is mediated by gustducin and TRPM5 (same as artificial sweetener).
3. Bitter Taste - Lots of different (26) receptors belonging to T2R family of GCPRs which create lots of heterodimer possibilities. Mainly mediated by TRPM5 and gustducin (same as artificial sweeteners and umami).
a. Phenylthiocarbamide (PTC) is tasted as bitter to 75% but not at all to the rest. This feature is autosomal recessive (variation in T2R38). PTC is similar to propylthiouracil (PROP), anti-thyroid drug that, along with PTC belongs to the thioureas class of drugs (because they contain N-C=S functional group).
b. Thioureas are found as 1-goitrin and isocyanates found in small amounts in broccoli, cabbage, Brusels sprouts, trunips, and kale. They will interfer with thyroid function in large quantities (inhibit iodine incorporation). Thioureas bitterness has been evolved to prevent excessive anti-thyroid drug action.
Cell structure:
1. Bipolar neurons with large number of dendrites (called chemosensory cilia, covered with receptors for odorants) that line the nasal mucosa and a single axon that synapse with cells of the olfactory bulb called mitral cells. Synaptic junctions between these two cells happen at glomeruli.
2. Each olfactory neuron only expresses a single odorant receptor gene. If a new odorant receptor gene is transfected into an olfactory neuron, the old odorant receptor gene is turned off.
3. Specificity of smells is defined by the combinations of olfactory neurons that are activated by a given odorant (as each odorant activates more than 1 neuron).
4. Olfactory neurons detecting the same odorant can be located anywhere in the nasal mucosa, but they all synpase in the same 1 or 2 glomeruli.
5. Glomeruli for similar odorants are located close to one another.
6. Some odorant receptor genes are expressedin testes and on spermatozoa.
1. Hydrophillic - Dissolve in the nasal mucous and interact with odorant receptors directly.
2. Hydrophobic - Interact with odorant binding proteins (OBP) found in mucous before binding to odorant receptors.
Receptor Structure/Mechanism:
1. G-protein-coupled receptor (GPCR), Golf, is a Gs type which activates cyclic-nucleotide gated channel (CNG, which lets in Ca++ which also opens a channel allowing Cl- efflux) through cAMP.
2. Note that Ca++ has a negative feedback effect on CNG to prevent uncontrolled feed-forward amplification of the odorant signal (called adaptation.
3. Extremely sensitive; a single odorant can activate 10 GPCRs resulting in 10,000 cAMP (only 3 are needed to open a CNG).
1. Anosmia (can't smell).
2. Kallman Syndrome - Anosmia (due to lack of olfactor bulb due to migrational problems) and hypogonadotropic hypogonadism.
Two types: Rods are highly sensitive at low light intensity and are most abundant. Cones are specialized to receive one of red, green, or blue and are mostly concentrated at the fovea.
1. Visual pigments (rhodopsins in rods encoded by RHO or OPN2 and not specifically named in cones but OPN1SW, OPN1MW, OPN1LW for short-wave (blue), medium wave (green) and long wave (red) respectively) densely populate the membrane structures of each photoreceptor cell. Consist of a G-protein-coupled receptor (GPCR) called opsin that is covalently conjugated to retinal (vitamin A aldehyde or 11-cis-retinal) at a lysine residue of the opsin through a Schiff-base linkage (double bond between C and N).
2. Upon exposure to light, retinal isomerizes to all-trans-retinal which dissociates it from opsin.
3. Opsin changes to activated form, called metarhodopsin.
4. Metarhodopsin binds to the G protein transducin which is a Gi,o, which loses a Talpha subunit. Metarhodopsin also is phosphorylated by rhodopsin kinase and this then bound by arrestin 1, preventing metarhodopsin from re-activating transducin.
5. Transducin minus Talpha binds to a cGMP specific phosphodiesterase called PDE6, which decreases cGMP concentration.
6. Decreased cGMP concentration leads to closure of CNG channels and hyperpolarization of the photoreceptor cells.
7. To return to baseline, transducin is reunited with Talpha subunit, which leads to increase of cGMP levels which opens CNGs to let Ca++ in, which inhibits receptor, G-protein, guanylate cyclase, and CNG channel itself. Also, all-trans-retinal binds to a non-opsin GPCR that converts it back to 11-cis-retinal, which reconjugates with opsin (which is dephosphylated) to form rhodopsin or color opsins.
8. Note that high intracellular levels of Ca++ activate recoverin which inhibits rhodopsin kinase which enchances rhodopsin regeneration.
Photoreceptor cell -> Brain
Recall that phototransduction results in hyperpolarization.
With no light:
1. Photoreceptors depolarize and secrete glutamate which inhibits bipolar cells with inhibitory glutamate receptors and stimulates those with excitatory glutamate receptors.
2. Bipolar cells with excitatory glutamate receptors in turn depolarize and secrete glutamate, stimulating OFF ganglion cells to send an OFF action potential to the brain.
With light:
1. Photoreceptors hyperpolarize and do not secrete glutamate, allowing bipolar cells with inhibitory glutamate receptors to depolarize but do not stimulate those with excitatory glutamate receptors.
2. Bipolar cells with inhibitory glutamate receptors in turn depolarize and secrete glutamate, stimulating ON ganglion cells to send an ON action potential to the brain.
Color vision defects
Caused by mutation in opsin genes. Xanopia = total loss of X vision. Xanomaly = decreased sensitivity to X.
Red = prot, Blue = trit, green = deuter.
All color vision defects: Congenital and non-progressive. Not considered a medical condition necessarily (no loss in quality of life).
Red-green color vision defects: X-linked recessive. Red and green opsins share 96% amino acid homology. Most of sensitivity between the two comes from 3 differences in amino acid codons. Caused by errors in homologous recombination, resulting in deletion and/or creation of a red-green fusion gene. Can be more sensitive to red, green, or both.
Blue vision defects: Autosomal.
Retinal PIgmentosa
Most common progressive disorder of retina. Generally adult onset.
1. Decreased night vision (nyctalopia) due to rod cell degeneration.
2. Narrowing of visual fields
3. Deteriorating color vision (cone cell involvement)
4. Complete blindness.
5. Characteristic patch areas of hypopigmentation adjacent to areas of hyperpigmentation.
Genetics: Some autosomal dominant, some autosomal recessive, some X-linked recessive, some sporadic.
1. RHO (for rhodpsin) -> can also cause a related disease, congential stationary night blindness.
2. PDE6A, PDE6B (for phosphodiesterase PDE6)
3. CNGA1, CNGA2 (for CNG channel)
4. RGR (for retinal GPCR)
Hearing (with genetic defects)
General steps:
1. Traveling wave created by stapes at the round window generates vibrations in the basilar membrane. Sounds of different frequencies result in vibrations of different intensitites along the length of the basilar membrane.
2. During the up and down movement of the basilar membrane, the positions of the hair cells move relative to the tectorial membrane.
3. The ends of each cilium are connected to the next cilium by molecular structures called tip links that ensure adjacent cilium deflect synchronously.
4. Deflection of the cilia towards the kinocilium triggers the opening of non-selective cation channels, resulting in depolarization and subsequent release of neurotransmitter.
1. Mutation of connexin 26 - Gap junction protein. 50% of autosomal recessive prelingual hearing loss is caused by this. Loss of gap junctions between hair cells impairs recirculation of K+ in inner ear and may disrupt cytoskeleton structure.
2. Tip Link problems - Myosin 7A, Cadherin 23, Protocadherin 15, and harmonin are required. Defects in genes for these proteins result in disorganization of hair cell bundles and deafness.
3. Usher Syndromes - Some of tip link genes are also needed for normal retinal development. So, this is characterized by severe congential hearing impairment and early onset retinitis pigmentosa.
4. Defect in tectorin - Extracellular matrix protein of tectorial membrane. Defect causes deafness.
5. Aminoglycoside-induced hearing loss - Mutation in 12S subunit of mitochondrial ribosome (so mitochondrial inheritance) results in tight binding of aminoglycoside and mitochondrial ribosome. So, no mitochondrial protein synthesis -> death of hair cells -> deafness. Also results in some renal toxicity but this is reversible due to renal cell regeneration.
Pain and temperature stimulation
Free nerve endings. Mediated by Transient Receptor Potential vanniloid subfamily (TRPV). Each TRPV has a different temperature set point. Heat-activated channels have open configuration while cold-activated channels have closed configuration. TRPV1 is activated by capsaicin which is found in hot red peppers.
Other factors:
1. Anandamide - Ethanolamide of arachidonic acid and acts upon these channels causing depolarization and thus pain and heat sensations. Also endogenous ligand for endocannabinoid receptors, which are GPCRs that cause hyperpolarization of sensory cells and thus cause pain relief and sedation.
2. Inflammation - Tissue hormone bradykinin, the neuropeptide substance P, and prostaglandin E can interact with corresponding GCPRs to activate phospholipase Cbeta which activates TPRV1.
3. Nerve Growth Factor - Strongly expressed in injured or inflamed tissues. Can bind trkA, a tyrosine kinase which activates phospholipase Cbeta which activates TPRV1.
1. Hereditary sensory and autonomic neuropathy type 4 (HSAN4) - Autosomal recessive. Mutation in gene for trkA. Patients cannot sense pain and so sustain injuries and infections. Also insensitivity to temperature and have poor wound healing. Also autonomic disfunction like anhydrosis (inability to sweat), postural hypotension, developmental delay, hyperactivity, and emotional lability. HSAN5 is caused by defect in NGFbeta.
2. Deactivating mutation in peripheral sodium channel SCN9A causes lack of pain, while activating mutation (autosomal dominant) causes priamry erythermalgia (PE) and paroxysmal extreme pain disorder (PEPD).
a. PE - Episodic burning pain, edema, and purplish or reddish discoloration of their distal extremities. Triggered by warm stimuli, exercise, or standing. Relieved by cooling or standing.
b. PEPD - 4 types of episode: Birth crisis (baby born red and stiff), Rectal crisis triggered by defecation and by emotional factors, Ocular crisis, Mandibular crisis triggered by eating and yawning. Also have skin flushing, reflex asystolic syncopal events, lacrimation, and rhinorrhea. Between episodes, patients are completely normal.
Bergmann's rule
Body size increases as climate becomes colder because as BM increases, the surface area doesn't increase proportionately, which changes the relative rate of heat loss in favor of energy concentration.
Allen's rule
Limbs become shorter as climate becomes colder because, as limbs becomes shorter , the surface area doesn't increase proportionately, which changes the relative rate of heat loss in favor of energy concentration.
Regulation of melanin synthesis
1. Proopiomelanocotin (POMC), a pituitary hormone precursor is cleaved to form alpha-melancyte stimulating hormone (alpha-MSH).
2. Alpha-MSH stimulates melanocortin receptor 1 (MCR1).
3. Step 2 can be inhibited by agouti signaling protein (ASIP).
4. Stimulation of MCR1 increases cytosolic cAMP, which promotes the maturation of melanin to form dark/brown eumelanins. Without MCR1 stimulation, maturation does not occur and the synthesis of pheomelanin is increased.
Harpending-Jenkins distance
(pi-pj)^2/(pa(1-pa)) where pi and pj are the allelic frequencies of any allele under consideration in population i and j, respectively, and pa is the average allelic frequency for that allele among all populations.
Phases of drug metabolism
Phase I: A functional group is inserted or activated on the drug substrate. Most common example is via cytochrome P450 enzyme (CYP) family. Can also include flavin-containing monooxygenases, hydroxylases, peroxidases, lipoxygenases, cyclooxygenases, monoamine oxidases, dioxygenases, reductases, and dehydrogenases. This can turn drugs into carcinogens, free radicals teratogens, or mutagens.
Phase II: A highly polar group is usually conjugated onto the phase I product. Examples are glucuronyl, sulfate, glutathione, acyl, amine, acetyl, and methyl-transferases.
Cytochrome P450 System (CYP)
Prototypic member of Phase I drug metabolism enzymes. Large family of enzymes that oxidize a wide variety of compounds using molecular oxygen. Use NADPH to provide ultimate reducing power in order to regenerate active enzymatic form with help of cytochrome P450 reductase. Localized in smooth ER. Most are involved in steroid and prostaglandin synthesis. Only CYP1, CYP2, and CYP3 are involved in drug metabolism.
N-Acetylation (NAT)
N-acetylases are prototypic examples of phase II drug metabolizing enzymes.
Example: Anti-tuberculosis drug isoniazid (INH) is metabolized (inactivated) by the enzyme N-acetylase. INH administration is associated with incidence of peripheral neuropathy in individuals with slow drug metabolism (and so high drug level). So, need to give INH and then measure INH levels 6 hours later to determine speed of metabolism.
Family studies: Slow acetylator parents always produce slow acetylator offspring but fast acetylator parents usually produce fast acetylators, but sometimes produce slow acetylators. This indicates that slow is recessive and fast is dominant. Slow acetylators is associated with increased breast cancer risk.
Thiopurine S-methyltransferase (TPMT)
Phase II drug metabolism enzymes that inactivates the chemotherapeutic agents 6-mercaptopurine (6-MP) and 6-thioguanin (6-TG). Slow metabolizing is recessive while fast is semi-dominant. Slow metabolizers given these meds have high complication rates from leucopenia and increased risk of radiation induced tumors and chemotherapy induced leukemia. This is directly related to overdosing the individuals based on their slow-metabolism.
Glucose-6-phosphate Dehydrogenase Deficiency (G6PD Deficiency)
X-linked recessive disorder. G6PD catalyzes the first step of the hexose monophosphate shunt, which is one of the major sources of NADPH. NADPH has a large role in the detoxification of hydrogen peroxide (2 pathways):
1. Glutathione peroxidase detoxifies H2O2 with the help of the reduced form of glutathione. NADPH restores glutathione to its reduced from after being oxidized by H2O2.
2. NADPH is a ligand for catalase, an enzyme that can reduce hydrogen peroxide to water.
1. Loss of reduction ability, which red blood cells are particularly susceptible to, which results in RBC membrane fragility and hemolysis.
2. Hemolytic crisis triggered by primaquine, sulfa, and fava beans.
3. If person is a slow-acetylator, then that can be synergistic with G6PD deficiency. For example, if exposed to analine. There are 3 ways to break down analine.
a. Acetylation of their amino group through NAT2.
b. Oxidization by the cytochrome P450 system to an arene oxide, which is highly reactive and binds covalently to cellular proteins and nuclear acids.
c. Arene oxides can be decomposed non-enzymatically or enzymatically through a glutathione-dependent process.
Note that a and c are dependent on acetylation and G6PD.
Alpha-1-Antitrypsin deficiency
Example of ecogenetic disease, since genetic defect predisposes individuals to common environmental toxins. Alpha-1-AT normally inhibits proteolytic enzymes, especially elastase. The uninhibited elastase destroys elastic tissue in the lungs causing emphysema.
1. Emphysema (increased incidence of COPD made way worse by smoking which oxidizes Alpha-1-AT active site and creates free radicals that do the same thing).
2. Can have cirrhosis of liver because a mutation can cause messed up folding of alpha-1-AT which induces apoptotic response.
Butyrylcholinesterase Deficiency (cholinesterase deficiency)
Autosomal recessive. Succinylcholine, a commonly used short-acting muscle relaxant for the induction phase in general anasthesia, is usually metabolized by butyrylcholinesterase. So, succinylcholine has a longer half-life in these patients, which means they will require mechanical ventilation for longer. Cocaine and procaine are also metabolized by this.
Autosomal recessive. Aka fish odor syndrome. Defect in gene for flavin-containing monoxygenase-3 (FMO3) develop a strong fishy odor after ingestion of food containing choline (present in fish, eggs, soybeans, and liver). Fishy odor is due to trimethylamine. Main problem is emotional and social problems.
Malignant Hyperthermia (MH)
Autosomal DOMINANT. Precipitated by a number of inhalation and muscle relaxing agents used in anasthesia. Symptoms:
1. Muscular rigidity
2. Extreme hyperthermia
3. Tachycardia
4. Increased oxygen consumption
5. Cyanosis
6. Hypercarbia
7. Respiratory and metabolic acidosis
8. Hyperkalemia
9. Myoglobinuria
10. Consumptive coagulopathy.
Treatment: Dantrolene
Mechanism: CAused by defect in ryanodine receptor (RYR1) which is a calcium release channel protein. This results in hypersensitivity to halothane-induced calcium release by sarcoplasmic reticulum in muscle cells.
Fetal isotretinoin effect (13-cis retinoic acid)
Affects neural crest cell migration, and prenatal exposure from the 2nd through 5th week post-conception results in craniofacial anomalies (small mandible, cleft palate, microtia, anotia) as well as thymic aplasia or hypoplasia, congenital heart disases (conotruncal and aortic arch defects) and brain anomalies (hydrocephalus, microcephaly).
Maternal PKU effects
Mother with poor control of PKU will expose fetus to high levels of phenylalanine. Causes low birth weight, microcephaly, mental retardation, and congenital heart disease.
Fetal alcohol effects
Causes growth and developmental delay, behavioral disorder, microcephaly, minor facial and skeletal anomalies and congenital heart disease. Least significant effects happen at 2 drinks a day, more subtle clinical features at 6 drinks a day. Most fetal alcohol syndrome babies are born to alcoholic women, but lesser effects can come from binge drinking or the like.
Hyperthermia fetal effects
Hyperthermia (fever > 38.9 for one day or more) during pregnancy has been associated with neural tube defects as well as clefting and a number of craniofacial anomalies. Can also be due to prolonged sauna or hot tub exposure.
Fetal Rubella Effect
When mother infected, 50% of time fetus is infected too. Causes intrauterine growth restriction, microcephaly, mental deficiency, deafness, cataracts, glaucoma, congenital heart disease, hepatomegaly, bone marrow suppression, and pneumonia.
Fetal Varicella Effects
1-2% of pregnancies w/ mothers w/ varicella results in infants with mental deficiency, seizure, microcephaly, chorioretinitis, limb hypoplasia, and cutaneous scars.
Regulation of heme synthesis
Bone marrow: Constant, controlled by erythropoitin and the availability of intracellular iron.
Liver: When excess heme, it is oxidized to hemin (Fe3+) and hemin decreases the transcription of ALA synthase, which is the major rate limiting enzyme in porphyrin synthesis. When drugs are present that require metabolism via cytochrome P450 system, there is an increase in cytochrome P450 enzymes. This decreases available heme, leading to an increase in ALA synthase and thus increase in heme production.
The Porphyrias
Group of 7 disorders resulting from defect in enzymes necessary for porphyrin synthesis (heme intermediate). Symptoms are due to buildup of intermediates in the heme pathway many of which are toxic.
1. Acute intermittent Porphyria (AIP) - Autosomal dominant. Reduction of porphobilinogen deaminase. Symptoms are latent prepubertally and manifestation is more common in females and are triggered by medication and/or hormone fluctuations. For example, barbituates induce synthesis of cytochrome P450 enzymes, which reduces pool of heme in liver. This results in loss of feedback on ALA synthetase which promotes flux through heme synthesis pathway. Reduction in PBG deaminase causes accumulation of aminolevulinic acid and porphobilinogen which cause neuro symptoms (including severe abdominal pain, tachycardia, hypertension, fever, confusion, hallucination, seizures, and other psychiatric symptoms).
2. Porphyria Cutanea Tarda (PCT) - Most common porphyria. Due to defect in uroporphyrinogen enzyme. Intermediates buildup in the skin, get radicalized by UV light, and damage the skin. Develops in 4th or 5th decade of life and triggered by hormones, alcoholism, iron overload, hep C, and HIV.
3. Lead poisoning - Lead inhibits ALA dehydratase and ferrochelatase resulting in block in heme synthesis. Results in increase in ALA and neuro symptoms similar to AIP. Decrease in heme production also leads to anemia.
Heme degradation
Begins in liver and spleen where senescent RBC's are taken up by macrophage. Heme oxygenase enzyme is 1st step which breaks down heme to CO2, Fe2+, and biliverdin. Biliverdin (which is green) is reduced to bilirubin (which is yellow). Same thing happens in bruises.
Bilirubin metabolism (including unconjugated hyperbilirubinemia)
Bilirubin is not soluble in plasma and so must be transported bound to albumin (in this state called unconjugated bilirubin). When reaches liver,
1. Enters hepatocytes and is conjugated in 2 step process by bilirubin glucuronlytransferase which adds 2 UDP moieties onto bilirubin making it more soluble in aqueous solutions.
2. Bilirubin diglucuronide is then excreted into bile (requires energy).
3. Bilirubin diglucuronide in small intestine is converted to urobilinogen by bacteria.
4. Most of the urobilinogen is oxidized to stercobilin (brown and gives stool its color).
5. Some urobilinogen is reabsorbed by the intestine and makes its way to the kidney where it is converted to urobilin (which is yellow and gives urine its color).
Lab Tests: Can measure direct bilirubin (bilirubin that is soluble aka conjugated bilirubin) and total bilirubin (all bilirubin in blood). Indirect bilirubin (unconjugated) is calculated by total - direct.
Juandice: Yellowing of skin and sclera due to bilirubin deposition.
Unconjugated Hyperbilirubinemia:
1. Hemolysis - Massive hemolysis resulting in overproduction of bilirubin can cause juandice. Liver's ability to conjugate bilirubin is overwhelmed so elevated unconjugated bilirubin in blood along with increased urobilinogen in urine.
2. Impaired hepatic uptake - Usually due to Gilbert's Syndrome (results in mild unconjugated hyperbilirubinemia during stress or times of fasting, not treated) or drug side effects.
3. Impaired conjugated - Due to Crigler-Najjar Syndrome. Decreased glucuronyl transferase enzyme. Serious disease presents early in life with jaundice and kernicterus (bilirubin deposits in brain) and results in death in a few years w/o treatment.
4. Benign Neonatal Jaundice - UDP-glucuronyl transferase takes about 4 weeks to reach full activity. Can be a problem because unconjugated bilirubinemia in neonates can diffuse across incompletely formed blood-brain barrier and lodge in basal ganglia. This is called kernicterus and can lead to toxic encephalopathy. Treatment is phototherapy which breaks bilirubin into urine-excretable products.
Conjugated Hyperbilirubinemia
1. Generally caused by block in bile duct (bile can't be excreted and backs up into the blood) or liver damage (causes increase in both conjugated and unconjugated bilirubin). Conjugated bilirubin is soluble in water and so some can be excreted via urine.
2. Dubin Johnson Syndrome is a defect in transporter that excretes bile into bile canaliculi. This increases conjugated bilirubin and blackening of the liver but isn't a big deal at all.
Principal protein for transfer of iron in the plasma and interstitial fluids. Synthesized in the liver. Has high affinity for Fe3+ (this renders Fe3+ soluble and prevents it from causing oxidative damage and facilitates iron transport into cells). Has 2 lobes and a Fe3+ is at center of each. An aspartate, a histidine, and two tyrosine residues for 4 and the remaining 2 coordinate bonds furnished by a bicarbonate ion. It is open when no Fe3+, but closes like a pincer if there is a Fe3+.
Lactoferrin has a similar structure and is found in neutrophils, secreatory epithelium and secretions such as breast milk. Has a much higher affinity for iron which makes it ineffective for transfer, but can sequester iron which people think makes it a good anti-bacterial.
Transferrin absorption to cells:
1. Transferrin receptors are found on the cell surface of erythroid and developing CNS cells. Consist of two glycoprotein homodimeric units connected by disulfide bonds.
2. Only transferrin with 2 iron bound have high affinity to transferrin receptors.
3. Upon binding to transferrin receptors, receptor-mediated endocytosis takes place via clathrin-coated pits.
4. ATP-dependent proton pumps on the vesicle acidify the vesicle to dissociate Fe3+ from transferrin.
5. Fe3+ (after being reduced to Fe2+ via a ferrireductase called STEAP3) is transported into cell via DMT1 transported that were on the part of the plasma membrane that was endocytosed.
6. The Fe2+ is transported into mitchondria for heme or Fe-S protein synthesis via mitoportin, or is taken up by cytosolic ferritin for storage.
7. Transferrin receptors-apotransferrin complex are (instead of being degraded in lysosomes) recycled back to the plasma membrane (apotransferrin is released extracellularly). This is called the transferrin cycle.
1. Iron response element (IRE) is a stem-loop structure that can be bound by a cytosolic protein called iron regulatory protein 1 (IRP1). IRE is present int he 5' UTR ferritin mRNA. (weirdly is similar to aconitase and has aconitase activity).
2. At low iron concentration, IRP1 binds to IRE, which impedes translation of the mRNA resulting in low ferritin subunit protein synthesis.
3. At high iron concentrations, a Fe-S complex binds to IRP1, making it unable to bind IRE allowing translation to proceed.
4. IRE is found in the mRNA of other protein involved in iron metabolism (including DMT1, ferroportin, mitochondrial aconitase, and some heme synthesis enzymes). For example, IRE is found in 3' UTR of transferrin receptors and, when bound by IRP, increases translation (so when high iron conc have low binding -> low translation but when low iron conc have high binidng -> high translation).
Intracellular iron storage (mainly in liver, spleen, and skeletal muscle).
Structure: Protein shell and core for iron (Fe3+) storage. Each molecule has 4500 iron atoms in a soluble, non-toxic, bioavailable form.
Synthesis: Initially called apoferritin, Fe2+ penetrates shell and is oxidized to Fe3+ via H subunit which causes nucleation and sustained growth of iron core.
Non-heme iron absorption (and regulation thereof)
Increased via ascorbate and citrate, decreased via fiber. Dietary iron is normally in Fe3+ and needs to be reduced to Fe2+ via ferrireductase for absorption. Fe2+ is transported into enterocytes via divalent metal transporter I (DMT1) which is a H+ symporter. Only works at low pH (also transports lead) and is upregulated with iron deficiency (a problem since iron deficiency can lead to lead poisoning).
Once in the enterocyte, Fe2+ can be used for enzymatic synthesis or stored in ferritin. Can also be exported through basolateral membrane through ferroportin (which is coupled to oxidation of Fe2+ to Fe3+ via haphaestin).
Hepcidin (a hepatic peptide hormone) regulates iron absorption by binding the ferroportin, internalizing and degrading them. Hepcidin release is increased during states of iron overload, inflammation and repressed during hypoxia and increased erythropoiesis.
Ceruloplasmin is involved in loading Fe3+ onto transferrin.
Iron Deficiency
1. Anemia
2. Impaired immunity
3. Decreased muscle and neurocognitive performance
4. Poor weight gain
5. Pica (ingestion of non-nutritious substances)
(more serious states of deficiency)
6. Epithelial changes in gastrointestinal tract including angular stomatitis, glossitis, papillary atrophy and nail deformity.
Hereditary hemochromatosis (HFE)
Autosomal recessive w/ very low penetrance. Most common disorder that predisposes to iron overload.
1. HFE gene malfunction results in low circulating levels of hepcidin, which leads a failure in the inhibition of iron export from enterocytes through ferroportin.
Symptoms: Tissue damage caused by iron overload.
1. Iron deposits in liver, heart, pancreas, endocrine gland, joints.
2. Hepatomegaly with fibrosis.
3. Congestive cardiomyopathy
4. Diabetes mellitus
5. Hypopituitarism
6. Arthritis
7. Cutaneous deposits resulting in hyperpigmentation (sometimes can have grayish hue or color).
Treatment: Therapeutic phlebotomy. No medicinal iron or large doses of vitamin C.
Iron poisoning
4 phases of clinical manifestation:
1. 30 min to 2 hrs: Acute abdominal pain, diarrhea and vomitting. CNS problems if high dose.
2. 6 to 24 hrs: Abatement of symptoms (NOT DONE).
3. 12 to 28 hrs: Metabolic acidosis, shock, interstinal perforation, hepatic and renal failure. Death.
4. 2 to 4 weeks: Abdominal obstruction secondary to intestinal scarring with multiple stricture formation.
1. Syrup of Ipecac to induce gastric emptying so less iron is absorbed. Only effective against pre-symptomatic patients.
2. Sometimes aggressive whole bowel irrigation with endoscopy.
3. Deferoxamin is an iron chelator that removes iron from transferrin and ferritin but not hemoglobin. Enhances urine excretion of iron (urine may become rose color).
Affects on hemoglobin by O2, CO2, H+, 2,3-DPG, CO
1. No oxygen - The 2 alpha-beta heterodimers are held together tightly through ionic and hydrogen bonds. This is called T (for "taut") configuration of the molecule because there is great constraint for movement of one heterodimer relative to the other. Extremely low affinity for O2.
2. Some oxygen bound - Positive allosteric effect, some of the inter-dimeric bonds are disrupted and the molecule assumes a more relaxed configuration called R. Affinity for oxygen goes up.
3. CO2 bound - Binds uncharged alpha-amino groups of the globin chains to form carbamino-hemoglobin. This stabilizes the T form which facilitates unloading of oxygen in peripheral tissues (SHIFT O2-BINDING CURVE OF HEMOGLOBIN TO RIGHT).
4. H+ bound - At peripheral tissue, RBC CO2 is converted to carbonic acid via carbonic anhydrase. Carbonic acid dissociates to H+ and bicarbonate. Deoxygemoglobin has higher affinity for H+ than oxygenated hemoglobin (Bohr Effect), thus H+ SHIFT O2-BINDING CURVE OF HEMOGLOBIN TO RIGHT.
5. 2,3-diphosphoglycerate (2,3-DPG) - Side product of glycolysis. Binds to central cavity in the hemoglobin tetramer via positively-charged amino acid residues in the globin chains, decreasing oxygen-binding affinity of hemoglobin and is useful for facilitating unloading of O2 in peripheral tissues (SHIFT O2-BINDING CURVE OF HEMOGLOBIN TO RIGHT).
6. Carbon monoxide (CO) - Extremely high affinity for hemoglobin (but reversibly binds). Converts hemoglobin to R configuration resulting in higher affinity for O2 (left shift). Since the CO-hemoglobin is not dissociated at peripheral tissues, the O2 is not effectively unloaded, causing hypoxia (patient have pink coloring due to high concentration of oxygenated hemoglobin in venous and capillary blood).
Hemoglobin F (as opposed to A1)
Higher affinity for oxygen than Hemoglobin A1 (left shift). Due to absence of some of the positively-charged amino acids residues in the gamma chains (compared to beta chains in A1) which are responsible for 2,3-diphosphoglycerate (2,3-DPG) binding. Thus, 2,3-DPG cannot right shift the curve. This is a special adaptation to the fetal environment when the partial pressure of O2 is substantially lower than in adults.
Sickle Cell Disease (Hemoglobin S)
Autosomal recessive. Mutation in hemoglobin Beta subunit making betaS (Bs). Heterozygous may be more resistant to malaria. In homozygous, mutation results in aberrant interaction in which Hb S polymerizes into a 14-straded rod-like helical structure at oxygen saturation less than 85% (40% for heterozygous) (which distorts the shape of RBC into a "sickle" configuration, increasing viscosity of blood). Note that oxyhemoglobin is more resistant to sickling (in other words, sicking is made worse by pH and high 2,3-DPG concentrations).
1. Increased viscosity of blood that causes vascular occlusion at small arteries and capillaries (this causes most complications of the disease).
2. Infection risk due to functional hyposplenism due to vaso-occlusive disease of the spleen. Mainly streptococcus pneumoniae and hemophilus influenzae.
3. Vaso-occlusive disease: Begins in infancy with dactylitius (hand-and-foot syndrome). Eventually get worse with periosteum involvement causing bone pain + fever. In adolescents and adults, involves lungs, kidneys, penis, liver, heart, and spleen. Can even increase risk for stroke.
4. Hemolytic anemia: Increased RBC fragility + vaso-occlusive disease greatly decreases RBC half-like through hemolysis. Can also cause gall-bladder disease.
Treatment: Gamma (y) subunits can replace mutated Beta (B) subunits. Hydroxyurea can increase expression of Gamma subunits.
Hemoglobin C
Amino acid substitution (missense) causes beta subunit to be betaC. This causes hemoglobin to crystallize (as opposed to polymerizing a la sickle cell disease), resulting in increased rigidity of the RBCs and consequently hemolytic anemia but no other symptoms.
MAJOR PROBLEM: When patients have BOTH Hb C AND Hb S (sickle cell disease). Can get really bad.
Hemoglobin E
Amino acid substitution (missense) causes beta subunit to be betaE. Homozygous is rarely symptomatic. Results in RBC microcytosis (decreased mean corpuscular volume).
The Thalassemias
Characterized by decrease in production of either alpha or beta globin chains, resulting in an imbalance.
a. Beta-thalassemia - Excess alpha-chain aggregates are insoluble, forming intracellular inclusions. This results in intramedullary hemolysis and hypersplenism, both of which lead to anemia.
b. Alpha-thalassemia - Intramedullary hemolysis is not a problem but hypersplenism remains. Splenectomy can help but cell destruction continues in liver.
c. Both - Continued stimulation of hematopoiesis results in expansion of bone marrow into ectopic sites leading to bone deformity and thinning with pathological fractures. Also, increased iron absorption couples with less use of iron = iron overload.
a. Alpha-thalassemia - If you have alpha-null (more common in African americans), alpha-alpha, you are good. If you have alpha-null, alpha-null, you might have minor symptoms but are still ok. If you have alpha-alpha, null-null (more common in southeast asia), you have minor symptoms again. If you have alpha-null, null-null you will have moderate to severe symptoms. IF you have null-null, null-null you are screwed (dead). In fetus you have gamma4 tetramers called Bart's Hb.
Increased proportion of blood volume taken up by RBCs. Majority of patient with this do not have globin gene mutations, but mutations that increase oxygen-affinity can cause this, as there is tissue hypoxia which stimulates hematopoiesis.
Polycythemia increases blood viscosity which can lead to a hypercoagulable state which can cause stroke or myocardial infarction.
Autosomal dominant. Due to oxidation of Fe++ to Fe+++ within the hemoglobin, thus stopping hemoglobin from binding O2.
1. Most commonly due to deficiency of cytochrome b5 reductase.
1. Chronic cyanosis with no pulmonary or cardiac issues.
2. Exacerbated by oxidizing drugs.
1. Can treat with methylene blue, except in patients with glucose-6-phophate dehydrogenase (G6PD) deficiency, since methylene blue-NADPH pathway is G6PD-dependent.
2. If G6PD deficiency, can use ascorbic acid (Vitamin C).
Vitamin A
General: Retinol (storage), Retinaldehyde (visual pigment), Retinoic Acid (hormone).
1. Vision - Cis-retinaldehyde absorbs photons which converts it to trans-retinaldehyde, triggering a shift in the structure of rhodopsin. Rhodopsin triggers an increased charge across the membrane in the rod and cone cells which blah blah vision.
2. Endocrine: Vitamin A is also converted to retinoic acid which interacts with cell via retinoic acid receptor. Additionally, vitamin A is fat soluble, membrane bound antioxidants.
3. Asorption/Storage:
a. Carotenoids are absorbed in enterocytes in the small intestine by passive diffusion and must be cleaved to form retinal. Supplemental retinol and beta-carotene are the most bioavailable.
b. Retinal is transferred to the liver as a retinal ester in chylomicrons.
c. In liver, retinol is bound to retinol binding protein (RBP) where it can be oxidized to retinoic acid.
d. Zinc is required to make RBP, so zinc deficiency can lead to Vitamin A deficiency.
4. Food: In meats, vitamin A is found preformed. In plants, found as beta-carotene.
5. Deficiency: Blindness, failure to thrive in children.
6. Toxicity: Increased skin pigmentation and increased risk of lung cancer. Liver damage + birth defects.
Vitamin B1 (Thiamine)
1. Precursor to Thiamine pyrophosphate (TPP), which is the cofactor for enzymes that add coenzyme-A to pyruvate and alpha-ketoglutarate. Necessary for carbohydrate metabolism, branched chain amino acid metabolism, and in the production of reduction equivalents.
2. Absorption: In stomach and small intestine. Alcohol interferes (so alcoholics = thiamine deficient often). Found in whole grains, nuts, and seeds. Gut bacteria also produce it.
3. Deficiency: Mainly in 3rd world and alcoholics. Limits activity of pentose phosphate shunt and subsequent loss of reducing equivalents can cause oxidative damage to cardiac and neural tissue.
a. Wernicke-Korsakoff encephalopathy - Acute deficiency. Characterized by anorexia, weight loss, mental status changes.
b. Chronic deficiency: Leads to beriberi. Wet form leads to heart failure + edema while dry form leads to peripheral neuropathy.
Riboflavin (Vitamin B2) and Niacin
Electron "Shuttle" Vitamins.
Riboflavin: Precursor to FAD and FMN which are primarily used as "electron shuttles" in redox reactions and converting folate and B6 into their active forms.
Niacin: Precursor to NAD which transports electrons to the electron transport chain. NAD is more negative redox potential than FAD which means it can't draw electrons from molecules as well as FAD, but it gives more ATP.
a. FAD - Limited absorption via alcohol, antacids, chelating agents (copper, zinc, caffeine, theophylline, vitamin C, tryptophan).
b. NAD - Can by synthesized from tryptophan and recycled by bacterial microflora in gut.
a. Mainly manifests in tissues with high levels of metabolic activity and/or high turnover (aka heart, musculature, skin, GI tract, and nerves).
b. Riboflavin = more mucous membranes (glossitis, red eyes, dry skin).
c. Niacin deficiency is also known as Pellegra and involves diarrhia, dermatitus, and dementia.
d. Hartnup's disease = Autosomal recessive where tryptophan is lost in urine and feces. Causes niacin deficiency.
e. Carcinoid syndrome = Loss of tryptophan due to serotonin producing tumor -> niacin deficiency.
f. Niacin can be used to increase HDL levels at the expense of vasodilation and flushing.
Vitamin B6 = Pyridoxine
Swaps out Amino acids. Precursor to pyridoxal phosphate (PLP), which is an electron sink for decarboxylation and deamination reactions necessary for the formation of amino acids, alpha-keo acids, and neurotransmitters. Also cofactor in rate limiting formation of gamma-ALA which is converted to heme and cytochrome products.
1. Unstable and is converted to PLP in liver upon absorption which requires zinc.
2. Alcohol inhibits absorption. Processing also destroys it (so processed foods).
3. Deficiency: Rare but leads to fatigue, dermatitis as well as a microcytic anemia (because of loss of heme production which leads to small, less pigmented red blood cells). Extreme cases, epileptiform convulsions can be seen. Some deficiency also contribute to mood disorders, pregnancy and amphetamine and cigarette usage.
Pantothenic Acid
Building C-C bonds. Precursor to coenzyme A. Here are some of what that does:
1. Solubilizes hydrophobic acyl chains in fatty acid synthesis
2. It provides a good leaving group during the formation of C-C bonds and is very important in fatty acid synthesis as well as the krebs cycle.
3. Plays a significant role in the acetylation of proteins such as histones.
4. Deficiency: Rare but disturbances in musculature (cramps), neural tissue (depression, paraesthesia, ataxia), and GI tract.
Decarboxylation. Carries CO2 and plays a limited role in the formation of malonyl-CoA from acetyl-CoA and in the conversion of pyruvate to oxaloacetate. Biotin is usually produced in excess by bacterial microflora in our gut.
Deficiency: Rare except when antibiotic usage and consumptions of craploads raw egg whites (which contains avidin, a substance that binds tightly to biotin and prevents its absorption). Symptoms = CNS depression and lethargy + dermatitis, conjunctivitis, and alopecia.
Folate and B12 (colambin)
Methylation exchange. Involved in homocysteine metabolism.
Folate: Helps build nucleotides, forming the purine ring which is necessary for coenzymes NAD, FAD, CoA and S-adenosylmethionine (SAM) which is the primary methyl group donor in the body.
1. Role of methylation - Needed for gluconeogenesis, amino and nucleic acids synthesis, neurotransmitter synthesis, myelin formation and regulation of the genome such as histone methylation and phase II of the liver detox pathway.
2. Folate absorption - Generally found in polyglutamate form, which in converted to a monoglutamate form prior to active transport in the jejunu. As a supplement it is unconjugated and more bioavailable. Heating can destroy it in food. Alcohol, methotrexate, and phenytoin can block its absorption.
3. Folate supplementation is encouraged in pregnancy because developing nervous system requires high levels of folate. If deficient, can have kid with neural tube defect.
4. Deficiency presents as macrocytic anemia. Some CNS effects can include weakness, fatigue, and difficulty with sustained attention, irritability, and headache + depression.
Vitamin B12 (colambin):
1. Has cobalt in it, which form corrin rings which must be synthesized by bacteria.
2. Utilizes a radical method, and is essential in methylation and 1 carbon transfers.
3. B12 binds factor-R in the saliva while intrinsic factor is released from the stomach. In the ileum pancreatic enzymes cleave factor R allowing intrinsic factor to bind colambin which allows it to be absorbed into the blood stream. Made more available by stomach acidity.
4. Present mainly in animal products (need supplementation for vegetarians).
5. Deficiency: Fairly common in elderly because low stomach acidity, poor diet, and gastritis all play a role. Characterized by demetia, sensory + motor disturbances, loss of memory disorientation. ANEMIA.
Vitamin C + E
Antioxidant Vitamins.
Vitamin C:
1. Antioxidant: Powerful in its own right and can regen Vitamin E and protect LDL particles from oxidation.
2. Collagen Synthesis: Vitamin C necessary to produce hydroxyproline from proline.
3. Absorption/Food Sources: Found only plant products. Excellent food sources = fruits and vegetables such as strawberries, bell peppers, broccli and brussel sprouts.
4. Deficiency: Causes scurvy, leading to deterioration of tissues which depend on collagen for structure (bleeding gums, impaired wound healing, joint pain, and easy bruising).
Vitamin E (tocopherols):
1. Lipid peroxidation: Mainly alpha-tocopherol. Associated with cell membranes. Stops LDL particles from being oxidized (recall this is a key step in atherosclerosis). Also stops pro-inflammation pathways (via protein kinase C and phospholipase A2).
2. Absorption/Food Sources: Requires bile salts. Incorporated chylomicrons and VLDL particles int eh liver and transported to adipose tissues for storgage. Low fat diets can limit absorption. In whole grains but not refined grains.
3. Deficiency: Peripheral neuropathy and sensory abnormalities (mainly in newborns and elderly). May cause oxidative damage in vascular and neural tissue.
4. Toxicity: Increase risk of hemorrhagic stroke and slightly reduce the risk of ischemic stroke. May involve inhibition of platelet aggregation or vitamin K.
Vitamin K
Assists blood clotting. Necessary for the carboxylation factors (II, VII, IX, and X) which can bind Ca2+ then form a blood clot.
1. Sources: Dairy products, nuts, beans, seeds, green leafy vegetables.
2. Hormones: PTH leads to increased levels of circulating calcium by releasing it from the bones, reducing calcium excretion by kidney, and by increasing dietary absorbtion of the calcium through activation of vitamin D.
3. Deficiency:
a. Causes: Lack of dietary calcium, low PTH hormone levels, vitamin D deficiency, hypoalbuminemia, chornic renal failure, and hypomagnesemia.
b. Symptoms: Paresthesias, tingling sensations, muscle spasms. Cardiac arrythmias, increased QT intervals. Osteoporosis.
4. Excess calcium: Generally indicative of underlying disease process.
a. Causes: Excess PTH due to parathyroid tumors, various cancers, chronic renal disease, disorders of Vitamin D metabolism, granulomatous diseases, and disorders of excess bone turnover.
Symptoms: "Painful bones, renal stones, abdominal groans, and psychic moans". Some symptoms caused by dehydration as kidney tries to excrete calcium (which requires water). This causes fatigue, nausea, and vomiting. Various neuro, muscular, and cardiac abnormalities appear (bradycardia, cardiac block, shortened QT, longer T wave).
1. Dietary Sources: Meat, dairy, grains (so most people are not deficient w/o disease process).
2. Regulation: Since it is contained in bone, involved with calcium metabolism and regulated by parathyroid hormone and Vitamin D. Absorbed in small intestine and filtered by kidney.
3. Deficiency:
a. Caused: Malabsorption (damage to GI system or deficiency of vitamin D), redistribution (refeeding of someone who is severely malnourished the enzyme phosphofructokinase becomes overactive), renal losses, alcoholism. PTH levels too high.
b. Symptoms: Disruptions of cell membranes and ATP defects lead to encephalopathy, dilated cardiomyopathy, generalized muscle weakness -> respiratory failure, hemolysis, rhabdomyolysis.
4. Excess:
a. Causes: Excess ingestion, laxatives, diabetic ketoacidosis, vitamin D toxicity, low PTH levels, disorders of cell lysis (like tumor lysis syndrome).
b. Symptoms: Increase in metastatic calcification in soft tissues due to formation of calcium phosphate crystals resulting in organ damage (especially kidneys) and also leads to hypocalcemia and all those symptoms.
Vitamin D
1. Sources: Skin exposure to light.
2. Mechanism:
a. Binds to Vitamin D Receptor (VDR), which dimerizes with RXR receptor and then binding to response element in the promoters of many different genes.
b. Increases of vitamin D lead to increased absorption of calcium in the intestine and increased deposition of bone.
3. Regulation: Exists in 2 main forms, Vitamin D3 (made by skin and in animals and best utilized by body) and Vitamin D2 (made by plants).
a. 7-dihydrocholesterol is converted to vitamin d3 by UV-B rays.
b. Vitamin D3 is converted into active form in liver or kidneys.
i. In liver, hydroxylate at the 25 carbon to form 25-OH D3 (calcidiol) which is main storage and circulating form.
ii. In kidneys, calcidiol is then hydroxylated either to inactive form 24-25 OH2 D3, or, if PTH present, converted to active form 1-25 OH2 D3 (calcitriol).
4. Deficiency: Decreased absorption of calcium and increased bone loss. Can be caused by kidney dysfunction, lack of sun, or hyopparathyroidism (not enough PTH).
a. in children: Rickets which is a defect in bone formation leading to soft pliable bones.
b. Adults: Osteomalacia, which is demineralization of bone leading to suseptibiltiy to fracture.
5. Excess: Increased calcium levels. Same as hypercalcemia. Sarcoidosis causes excess vitamin D via activation of D via macrophages.
Type of electrical or osmotic ion.
1. Biochem: Main extracellular ion (maintains electrical and osmotic balance in a cell.
2. Deficiency (too little Na outside cells): Leads to CNS edema causing nausea, vomiting, headache, confusion, irritability, fatigue, hallucinations, and muscle weakness.
3. Excess: Due to over-consumption (generally of processed foods). Long run kidneys don't regulate Na well, which can lead to hypertension.
Type of electrical or osmotic ion.
1. Biochem: Essential for maintaining osmotic potential (it is the intracellular cation). K+ flux is important for action potentials. K+ is pumped out of kidney in response to aldosterone as well as decreases in pH. It is also drawn into cells by insulin.
2. Deficiency:
a. Primary - Found in fruits and veggies. Leads to cardiac arrhythmias and muscle weakness (due to inability to maintain proper potential across the cell) as well as glucose intolerance (because K+ is needed for insulin release).
b. Subclinical deficiency - Exacerbate hypertension (although Na+/K+ ratio is more closely tied to htn than the level of either alone).
Type of electrical or osmotic ion.
1. Biochem: Primary anion in most cellular reactions (especially important in acid-base balance). Utilized as part of HCl in stomach.
2. Deficiency: Rare but can cause growth failure, lethargy, irritability, weakness, and GI distress.
Type of Bone mineral.
1. Biochemistry: Thought to be a "chronic regulator" of biological functions. Cell growth, energy metabolism, and cell death. Needed for carb metabolism, fatty acid and protein synthesis.
a. Energetic reactions: ATP(2-) is stabilized by complexing with magnesium. Mitchondria have large magnesium and ATP stores. So, anything that requires ATP also requires magnesium.
b. DNA and the cell cycle: Magnesium binds phosphate in the phosphate linkages of DNA and RNA, stabilizing the molecule for reactions. Magnesium depletion stops cell cycle by regulating cell cycle inhibitors p21, p27, cyclins and CDKs.
c. Glutamate receptors: Magnesium blocks glutamate-NMDA receptors in neural tissue creating a voltage sensitive block which prevents downstream neural activity. Depolarization or phosphorylation is necessary to remove the magnesium.
d. Muscle contraction: Inhibits the cross-bridge cycle as it binds the same spot as calcium.
2. Absorption: Large intestine (passively). Kidneys regulate excretion (magnesium is utilized to form urea).
3. Food: Centrally bound to chlorophyll (like iron in heme). Green = magnesium.
4. Deficiency: Neuromuscular excitability and other things that look like calcium deficiency. Also impaired insulin excretion.
5. Excess: Diarrhea. Drowsiness, lethargy, weakness. Antacids and laxatives have Mg, which can cause excess.
Like zinc, found mainly in coordination complexes. Absorbed in upper small intestine and share a transporter with Ca. Excess amounts of zinc or Ca can limit copper absorption. Shellfish has a lot (as well as zinc) as do whole grains and legumes.
1. Biochem:
a. Oxidase enzymes: Copper functions in several oxidase enzymes, including cytochrome oxidase, in the electron transport chain, super-oxide dismutase, a free radical scavenging enzyme in the immune response, lsyl oxidase, necessary for collagen synthesis and monamine oxidase, which functions in neurotransmitter catabolism.
b. Ceruloplasmin: In the serum, copper is found bound primarily to ceruloplasmin. In addition to mobilizing transoprt of iron into RBCs, ceruloplasmin is an acute phase reactant in the immune response, a compound which is produced in response to inflammation.
2. Deficiency: Rare mainly seen in kids with Menkes disease.
a. Menkes disease: X-linked recessive. Alters bodies distribution of copper. Children are identified because hair is extra-kinky. Causes skeletal abnormalities, failure to thrive, and recurrent infections.
b. Dietary deficiency: Mainly seen in gastric bypass surgery and excess zinc. Neuro symptoms result, including ataxia and neuropathy. Can also cause anemia due to ceruloplasmin requirement for iron to be incorporated into RBCs. Subclinical can cause recurrent infections.
3. Excess: Only seen with very high levels of supplementation, ingestion of fungicides, or contaminated water. Mild = GI distress. Major = severe brain and liver damage.
Like copper, found mainly in coordination complexes. Absorbed in upper small intestine and share a transporter with Ca. Shellfish has a lot (as well as copper) as do whole grains and legumes.
1. Biochem:
a. Enzyme cofactor: Lots of enzymes, including carbonic anhydrase (interconverts CO2 and bicarbonate) and DNA and RNA polymerases.
b. Zinc fingers: Transcription factors require specific geometric conformation to bind DNA. Zinc binds amino acids of an alpha helix and antiparallel beta sheet to structurally define the zinc finger.
c. Prostate gland function: Normally, prostatic epithelial cells secrete high levels of citrate for seminal fluids. Zinc regulates m-aconitase, which turns citrate into isocitrate. Too much zinc can therefore prevent prostate from secreting citrate.
d. Brain function: Released into synapses and therefore modulates synaptic transmission and plasticity. Excess causes neural excitotoxicity -> neural and glial cell death.
2. Deficiency: Common with diabetes, alcoholism, and liver diseases.
a. Symptoms: Hypogonadism, deformed bones, poor wound healing, abnormal hair or nails, loss of taste and CNS abnormalities.
b. Acrodermatitis enteropathica - Seen in infants. Absorption and transport of zinc is impaired and causes pustular or bullous dermatitis, growth retardation, failure to thrive, and secondary infections. Cured with oral therapy.
3. Excess: Not typically seen. Metal fume fever presents with flu like symptoms for several days.
Insulin receptor mineral.
1. Biochemistry: Component of apo-chromodulin = enhances effects of insulin. So, can give chromium to help with insulin resistance.
2. Excess: Can cause cancer.
Mineral for gultathione and thyroid function.
1. Biochem:
a. Antioxidant enzyme: Component of glutathione peroxidase, the principle intracellular antioxidant enzyme.
b. Thyroid function: Binds to de-iodinase enzymes which link iodine to thryoid hormones.
2. Deficiency:
a. Keshan's Disease - Causes cardiac and muscular wasting. Likely due to inhibition of glutathione production and thus inability of tissue to clear free radical. Generally found in countries with low levels of selenium in soil.
3. Excess: Reduces lung, stomach, colorectal, and prostate cancers with an increase in skin cancer.
Necessary for the formation of T3 and T4, the thyroid hormones. Salt is now iodinized. Can be absorbed through the skin (esp via automobile exhaust).
Creatine Formation and Utilization
Generation of phosphocreatine:
1. In kidney, guanidinoacetate is formed from glycine and arginine.
2. In liver, it is methylated to form creatine.
3. In muscle cells, it is phosphorylated via creatine kinase to form phosphocreatine.
1. Phosphocreatine generates ATP by donating P to ADP to form ATP and creatine.
2. Can also be converted to creatinine which is metabolically inactive but is used as a marker for renal function.
Supplementation: Good for short burst exercise but not for endurance. No real risks @ 3g/day (liver and kidney problems have been debunked).
Whey Protein
Contain branched chain amino acids (BCAAs, leucine, isoleucine, and valine). These initiate protein synthesis. Leucine can activate mTOR which phosphoylates targes taht upregulate transcription factors that initiate translation.
1. Faster recovery.
2. More muscle growth.
Downsides: GI problems, lactose intolerance can be an issue.
Types of muscle fibers
3 types:
1. Type I - Aka slow-twitch or red muscle (dark meat). Resistant to fatigue, has abundant mitochondria, increased capillary density, and relies on fat oxidation as major fuel source.
2. Type IIx (IIb) - Aka fast-twitch or white muscle (white meat). Quicker to fatigue, sparse mitochondria, utilizes glycolysis for ATP production.
3. Type IIa - Considered fast-twitch but really an intermediate fiber with ability to utilize glucose or fat for energy.
Effects of endurance exercise
1. Release of catecholamines (epinephrine and norepinephrine) and thus reduced basal rates of these = lower heart rate/blood pressure.
2. Insulin sensitivity (good for type II diabetes prevention!!!)
3. Increases mitochondria density (mediated through NGF1/2 (control expression of mitochondrial-derived genes in electron transport chain) and early growth response gene-1 (EGR-1) (increases expression of oxidative phosphorylation protein cytochrome c oxidase (COX).
4. Peroxisome proliferator-activated receptor (PPAR)-gamma co-activator 1alpha (PGC1alpha) -
a. Upregulates NGF1 (which has effects as above).
b. Increases expression of PPARs. This increases lipid trafficking, beta-oxidation, and uncoupling of the electron transport chain.
c. Activates mitofusin - Regulates mitochondria fusion and fission to promote continued mitochondrial efficiency.
d. PGC1alpha is itself activated via Calmodulin dependant kinase (CamK, which is activated via calcium) and AMP-activated protein kinase (AMPK, activated via AMP).
Resistance exercise
Two mechanism for muscle hypertrophy:
1. Increases efficiency of translation from existing mRNA, which is mediated by phosphatidylinositol 3 kinase (PI3K) - Akt (PKB), which activates the mTOR pathway. mTOR integrates numerous cellular signals such as those for enzyme activity, cellular redox status, and nutrient levels to modify cellular protein synthesis. Activates Elongation factor 2.
2. Activates skeletal muscle satellite cells, which appears to be primarily mediated by the damage caused during eccentric contractions (slowly letting down a muscle). Neighboring satellite cells, located in the basal lamina of a muscle fiber, are activated to reapir the muscle and increase myonuclei. Myogenic regulatory factors (MRFs), through myogenic differentiation factor (MyoD) and myogenin (MyoG) which act as transcription factors.
Affect of concurrent resistance and endurance training
They are not as efficient as either alone. Two hypotheses:
1. CaMK (from endurance) inhibits elongation factor 2 (EF2) while mTOR pathway activates EF2.
2. AMPK (from endurance) inhibits mTOR pathway, inhibiting protein synthesis in favor of mitochondrial biosynthesis.
Angiotensin-converting enzyme (ACE) polymorphism
Example of performance enhancing polymorphism. I allele has an insertion that is associated with decreased enzyme activity while D is normal. People who have I/I allele get more efficient muscle after endurance training as opposed to D/D. Thought to be because since there's less ACE activity, more NO release and increased efficiency of mitochondria.
alpha-actinin-3 (ACTN3)
Example of performance enhancing polymorphism. Aids in production of Z-line within sarcomere. Also coordinates myofiber contraction during actin-myosin crossbridge cycle. There's a normal protein and a null mutation (577X). In sprinters, significant increase in normal/normal genotype. In endurance, increase in null/null genotype. Could be because this is mainly seen in fast twitch fibers.
Myostatin (MSTN)
Example of performance enhancing polymorphism. Negatively regulates muscle growth and hypertrophy. Loss of this means more muscle mass. Mechanism is that it usually activates p21, which arrests cell cycle, so lack of MSTN = muscle hyperplasia not hypertrophy.