Gene Expression III

Question 1

How do you regulate SSDBPs?

Answer 1

1. ) Can alter the conformation of the DNA-BP via ligand binding
2. ) Can regulate its entry into the nucleus
3. ) Can control the amt of trx factor in the cell
4. ) DNA binding can be regulated
5. ) Phosphorylation of the DNA-bp can alter various properties including protein degradation, recruitment of co-activators, and DNA binding

Question 2

Ligand binding

Answer 2

An example is when androgen/estrogen bind the nuclear receptor family of Zinc finger trx. factors… which affects the dimerization, recruitment of coactivators/repressors, and translocation into the nucleus

Question 3

Tamoxifen

Answer 3

This is an estrogen antagonist that binds Zinc finger nuclear receptor trx. factors, so they cannot dimerize. So you can’t get proliferation etc.
Prevents HAT recruitment. may recruit co-repressors.

Question 4

NF-KB (example of SSDBP regulated by nuclear entry)

Answer 4

NOrmally held in cytoplasm through its binding to inhibitor IkB (it masks the nuclear localization signal).
IKB can get phosphorylated and degraded… so NKB goes to the nucleus and turns on target genes involved in inflammation.
Aspirin inhibits this phosphorylation so NF-KB stays in the nucleus!

Question 5

NF-AT (example of SSDBP regulated by nuclear entry)

Answer 5

High intracell Ca (from ligand binding to extracell receptor) activates calcineurin’s phosphatase activity
It dephosphorylates NF-AT, exposes its nuclear localization sequence, and it goes to enter the nucleus and affect trx of genes for immune response and heart fx!

2 common immunosuppressants INHIBIT CALCINEURIN so thus inhibit NF-AT action.

Question 6

Beta-catenin (example of regulating amt. of trx factor in the cell)

Answer 6

In absence of Wnt signalling, cytoplasmic pool of B-catenin is targeted for degradation via the ubiquitin/proteosome pathway (through phosphorylation with glycogen synthase 3, GSK3) in a complex with Axin and APC.

In presence of Wnt the Axin-APC-GSK3 complex is not stable. So B-catenin not phosphorylated and amount in cytoplasm increases. So some b-catenin moves to nucleus.
Where it interacts with TCF family of trx factors and promotes expression of Wnt responsive genes

COLON CANCER: ppl with polyps have mutations in APC (which allow b-catenin stabilization, and ultimately get trx of genes that cause proliferation….cancer)

Question 7

P53 (example of regulating amt. of trx factor in the cell)

Answer 7

Downregulated by binding of MDM2 protein (masks its activation domain and targets it for destruction) via the ubiquitin- proteosome pathway

A trx factor, activates cell cycle arrest genes and cell death processes (guardian of genome)

Question 8

Mechanism of inhibiting DNA binding activity of an SSDBP?

Answer 8

Id proteins: they heterodimerize with other HLH proteins through their HLH domains and prevent DNA binding due to their lack of a basic domain

Question 9

How does phosphorylation affect trans-acting factors?

Answer 9

CREB (cyclic AMP response element binding protein)- ligand binds to guanine nucleotide binding protein coupled receptor… get Phosphorylation of CREB (which while on DNA is inactive to promote trx. unless phosphorylated)

Once activated it recruits the HAT CBP (creb binding protein) which has HAT activity and recruits RNA POL II… get trx activation.

Question 10

Other ways to control levels of gene expression?

Answer 10

1. ) Control mRNA export from nucleus
2. ) Control mRNA degradation
3. ) Control efficiency of translation
4. ) Control of protein degradation