Regulation of Transcription by RNAPII

Question 1

What is the galactose utilization pathway in yeast? when is it on or off

Answer 1

• ex. of short-term transcriptional regulation
• 3 genes: GAL1, GAL10, GAL7
• turns B-galactose into G1P for glycolysis
• genes are only activated when galactose present and glucose absent
• otherwise repressed

Question 2

What does the genes look like for GAL1, 7, and 10?

Answer 2

• GAL 1 and 10 share a divergent promoter
• GAL 7 has its own promoter
• there are 2 consensus sequence protein binding domains called UAS(G)
• ## GAL1 gene has 4 of 17 mer sites bindings sites

Question 3

What is the Gal4 class?

What happens when mutated?

Answer 3

• protein that binds to UAS(G) sites in promoter
• transcriptional activator homodimer

when mutated, GAL1, GAL7, GAL10 are not synthesizes whether or not galactose is there

Question 4

What is the Gal80 class?

What happens when mutated?

Answer 4

• galactose absent: binds to Gal4 activation domain to block activation
• galactose is present: galactose binds to Gal3, which bind to Gal80, stopping Gal4 inhibition

when mutated, costitutive expression of all 3 genes

Question 5

What is the Herpes Simple Virus Transactivator V16?

Answer 5

example of a regulatory switch and model of combinatorial control (HSV-1 virus)

Question 6

What are the 3 classes of genes in HSV-1?

Answer 6

1) immediate early: regulatory functions
- small, early amount that drops off

2) early: DNA synthesis (replication) and nucleotide metabolism
- larger amount that increases and drops off after

3) late: structural components
- huge amount that increases a lot

interested in what causes this cycle

Question 7

What is VP16? What does it do and how does it work?

Answer 7

• protein that activates immediate early gene transcription of HSV-1 virus
• can’t bind to DNA on its own

IE promoter has TAATGARAT elements upstream, which Oct-1 and HCF1 binds

Question 8

What is HCF-1?

Answer 8

host cell factor 1
- protein required to deliever VP16 to Oct1 bound DNA
- forms larg complex

Question 9

How does the casade of immediate early genes transcription occur?

Answer 9

1. VP16 binds to HCT-1
2. Oct-1 binds to TAATGARAT sequence in upstream region of IE promoter
3. VP16-HCT1 binds to Oct1
4. gene expression
5. DNA replication
6. infection

Question 10

What are the functional domains of V16?

Answer 10

• core (part that binds to Oct-1 and HCF-1)
• TAD: mediator, TFs bind

transcriptional activation domain

Question 11

What is Oct-1?

Answer 11

• binds to TAATGARAT in UAS of IE promoter
• cellular factor
• member of POU homeodomain family of TFs (has POU domain)

POU domain has a POU specific domain and homeodomain

Question 12

What are the minimals domains for each protein in the IE cascade required for VP-16 complex formation?

Answer 12

VP - Core
Oct-1 - POU domain
HCF-1 - VIC domain

Question 13

How is combinatorial control of transcription by the VP16-induced complex shown?

Answer 13

1. selective recruitment of Oct-1 (not Oct-2)
2. both HCF-1 and HCF-2 stabilizes VP16-complex but only HCF-1 activates transcription effectively

VP16 discriminates with combinatorial interactions to drive transcription (makes combinations of different types of Oct and HCF types)