L4, Activators, Repressor and Chromatin in Euk. Gene Expression

Question 1

Where do activators bind? Implication for activator structure and function?

Answer 1

• Sequence specific enhancer or silencer regions
• An activator thus requires two key domains: one to bind the DNA and one to activate the transcription -> confers two different levels of specificity
• The activation domain function can be carried out by a separate coactivator/repressor

Question 2

3 common structures of sequence recognition in DNA binding domain of actviators:

Answer 2

• Zinc finger
• Helix-turn-helix
• Leucine Zipper

Question 3

Where are enhancer/silencer sequence located; how is this permitted?

Answer 3

• Many are far away from promoter either upstream or downstream
• If they are far away enough, the mediator complex will be required to loop the DNA into proximity with the general TFs
• Various other cofactors play specific roles

Question 4

What is the mediator complex?

Answer 4

• Aka coactivator
• Multisubunit complex which functions within the pre-initiation complex, and brings activators into proximity with basal TFs
• Size: 4 mega daltons (Huge!)
• Able to regulate basal TF activity (not fully understood)
• Structure comprises a head, body and tail region (head at basal TFs, tail at response element)

Question 5

Additional functions of mediator complex:

Answer 5

• Beyond DNA loop interactions, it is able to interact with ncRNAs
• Interacting with various regulatory factors -> epigenetics, elongation, termination, splicing etc (not just transcription)

Question 6

What are superenhancers?

Answer 6

• Region in mammalian genome comprised of multiple enhancer elements
• Bound by an array of TFs -> particularly high level of mediator binding
• Likely to be important for tissue specific gene activation etc (birds in hats analogy) -> different combinations of components conveying huge levels of specificity
• Good example of cooperative control

Question 7

List 5 ways that a transcription regulator can be modulated:

Answer 7

• Protein synthesis e.g. inhibitor
• Ligand binding
• Covalent modification (phosphorylation)
• Addition of second subunit
• Stimulation of nuclear entry

Question 8

What are insulators/barrier elements?

Answer 8

• Control action of activators; physically block a gene from indiscriminate activation / repression
• Act to divide up the genome into domains
• They can block enhancer and promoter regions
• They can also block spreading of chromatin modifications from one domain to another

Question 9

Rough roles of chromatin:

Answer 9

• Packaging genetic material into nucleoid/nucleus of cell
• Regulating gene expression and DNA replication
• Preventing DNA damage. regulating DNA repair etc

Question 10

+ Archaeal histone-based chromatin:

Answer 10

• Form a structure similar to the eukaryotic tetramer-based arrangement
• Arranged around histones in an ‘endless nucleosome’

Question 11

What two main modes can be used to alter chromatin state:

Answer 11

• Chemical modifications e.g HATs/HDACs
• Remodelling using chromatin remodelling complex (SWI/SNF); energy dependent displacement which occurs in conjunction with activation of genes

Question 12

What histone variant might be apparent at sites of DNA damage?

Answer 12

• H2AX

Question 13

Chromatin remodelling complex in higher eukaryotes:

Answer 13

• Evolved past SWI/SNF to more complex version
• Most recently (evolutionary time): nBAF

Question 14

What are transcription factories?

Answer 14

• Particular foci in nucleus
• Enriched for RNA PII
• Hotbeds of transcription
• Allows for spatial level of control