Chapter 17 Transcriptional Regulation in Eukaryotes

Eukaryotic genes are controlled by DNA sequences

1. Promoter (TATA box and Inr/initiator)

2. Promoter-proximal elements (GC-rich box ~-100 and CCAAT box ~-80)

3. Regulatory elements (enhancers and silencers) also known as ________________.

4. Insulators
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Eukaryotic genes are controlled by DNA sequences

1. Promoter (TATA box and Inr/initiator)

2. Promoter-proximal elements (GC-rich box ~-100 and CCAAT box ~-80)

3. Regulatory elements (enhancers and silencers) also known as ________________.

4. Insulators
3. Regulatory elements (enhancers and silencers) also known as response elements
2. Promoter-proximal elements (GC-rich box and CCAAT box)
very important in transcription
3. Regulatory elements (enhancers and silencers) also known as response elements effects and location can be investigated by:
reporter gene constructs
Reporter Gene Constructs
can be enhancer or silencer; can be up/downstream even 1000s of basepairs away can be in any orientation and can even be in the gene itself

you add on a gene that has products that can be measured and quantified ex : GFP
PQ: what is the difference between the Inr (initiator element) and the TATA box?
RNAP binds to the TATA sequence and then initiates transcription downstream at Inr.
PQ: An enhancer sequence.....
can be inverted without altering its functional ability
A ____________ DNA sequence can be located thousands of nucleotides away from the genes they regulate
enhancers
Whereas GTFs are necessary for basal transcription of all genes, RTFs control the rate of transcription of target genes.

RTFs recognize and bind to specific DNA sequences called regulatory or response elements (REs).

RTF amino acid residues and DNA nucleotides interact in the __________ groove of the DNA double helix via: H-Bonds, ionic bonds, hydrophobic interactions.
major groove
RTFs possess one or more of the following domains:
1. DNA-binding domain (DBD) - binds to a regulatory element
2. Activation domain (AD) - binds to RNA Pol II and/or GTFs
3. Transactivation domain (TAD) - binds to other RTFs
...
RTFs can be modulated by:
binding of a small effector molecule like a hormone,

Protein- protein interaction

covalent modification such as phosphorylation
GCR is an activator modulated by: 1. 2.1.) hormone binding 2.) dimerizationNLS = GRE =NLS = nuclear localization sequence GRE = GC response elementCREB is an activator modulated by: 1. 2.CREB is an activator modulated by: 1. Dimerization 2. PhosphorylationCRE = CREB =CRE = cAMP response element CREB = CRE-binding proteinHIF-1 is an activator modulated by 1.) 2.)HIF-1 is an activator modulated by: 1. Phosphorylation 2. DimerizationHIF-1 (cont.) von Hippel-Lindau disease (AD) constant upregulation of proteins and genes that lead to the formation of new blood vesselsLeads to hemangioblastomas tumorsGRE, CRE, HRE are ..........enhancers and RTF binding sitesGCR, CREB and HIF-1 are al modulated by:dimerizationFunctional genomics is the study of the function, expression, and interaction of gene products. "omics" include:"Omics" include: transcriptome = the sequence and expression patterns of all RNA transcripts (ex: which kinds, where, when, how much) proteome = the sequence and expression patterns of all proteins interactome = the complete set of physical interactions between proteins and DNA segments, between proteins and RNA segments, and between proteinsUsing DNA microarrays to study the transcriptome Question: Which genes are active in a particular cell under certain conditions?ex: period of development, exposure to a drug, etc.Using the yeast two-hybrid system to study the protein-protein interactome Question: Do proteins A and B interact?a protein protein interaction will result in expression of a reporter geneUsing the ChIP (chromatin immunoprecipitation) assay to study the protein-DNA interactome Question: What DNA sequence does a certain RTF bind to?studies protein dna interactomeUsing the band shift assay to study the protein-DNA interactome Question: What DNA sequence does a certain RTF bind to?studies protein DNA interactomeUsing the DNase footprinting assay to study the protein-DNA interactome Question: What DNA sequence does a certain RTF bind to?studies protein DNA interactomePQ: Which of the following would not be used to study the interaction between an RTF and an enhancer?DNA microarrayTwo modular elements that appear as consensus sequences upstream from RNA polymerase II transcription start sites are ________.TATA and CAATWhich of the following clusters of terms applies when addressing enhancers or silencers as elements associated with eukaryotic genetic regulation?cis-acting, variable orientation, variable positionthree characteristics of enhancers and silencers :They can regulate other genes when inserted. Their orientation may be inverted without significant effect. They can act at a great distance from the promoter.Mutations in the promoter region of the β-globin gene indicate that some areas are more sensitive than others. When mutations occur in consensus sequences (modular elements such as GC box, CAAT box, and TATA box), does transcription usually increase or decrease?Transcription should decrease because by changing these modular elements, the general transcription factors cannot bind efficiently or recruit RNA polymerase.Which of the following elements function specifically in eukaryotic transcription and gene expression?promoters and enhancers Promoters are the sites of assembly of the basic transcription complex, and along with enhancers, control transcription in eukaryotes.Transcription factors are important molecules that regulate gene activity in eukaryotes. What are the two general classes of transcription factors that exist in eukaryotes?general transcription factors activators and repressorsWhere do they bind and what are their effects? 1.) General transcription factors: 2.) Activators and repressors:1.) General transcription factors: assemble at promoter regions adjacent to the site of transcription important in basal levels of transcription 2.) Activators and repressors: are important in tissue specific and developmental regulation are responsible for altered transcription bind at more distant regions, such as enhancers and silencer sequencesWhy must a transcription factor have at least two functional domains?One domain binds to DNA in promoters and enhancers, and the other binds to other transcription factors or RNA polymerase to activate transcription.