Question

(1) Answer the three questions for each of the following transcription factors:

LacR

TrpR

CAP

GAL4

GR (glucocorticoid receptor)

(a) What is the small molecule that controls its’ activity?

(b) In the presence of this molecule, is the protein on the DNA or off the DNA?

(c) What is the consequence for transcription?

(2) Fill out the table with the words ON or OFF:

	- lactose: lac repressor ON or OFF the DNA	- lactose: lacZ ON or OFF	+ lactose: lac repressor ON or OFF the DNA	+ lactose: lacZ ON or OFF
I+ O+ Z+
I- O+ Z+
I+ Oc Z+
I+ O+ Z+ / Oc
I+ O+ Z+ / I-
I- O+ Z+ / I+
I+ Oc Z+ / O+

(3) Answer the three questions for each of the following enzymes:

Swi/Snf

HAT

HDAC

DNMT

(a) Describe the activity of each of the enzymes.

(b) What is the consequence of this enzyme activity?

(c) How does this enzyme know what genes to act on?

Answer 1

2)

	- lactose:lac repressor ON or OFF the DNA	- lactose: lacZ ON or OFF	+ lactose:lac repressor ON or OFF the DNA	+ lactose: lacZ ON or OFF
I+ O+ Z+	On	Off	Off	On
I- O+ Z+	Off	ON	OFF	ON
I+ Oc Z+	ON	ON	Off	ON
I+ O+ Z+ / Oc	ON	ON	OFF	ON
I+ O+ Z+ / I-	ON	ON	OFF	ON
I- O+ Z+ / I+	ON	ON	OFF	ON
I+ Oc Z+ / O+	ON	On	OFF	ON

3) a) Activity of each enzymes -

• Swi/Snf (Switching/ Sucrose Non Fermenting - SNF) - SWI/SNF is a nucleosome remodeling complex found in both eukaryotes and prokaryotes. They use the energy of ATP hydrolysis to remodel nucleosomes and to modulate transcription.
• HAT (Histone acetyl transferase) - It is a histone modification enzyme, that catalyze the addition of acetyl groups to the lysines of the histone N-termini. Their are different HAT enzymes that target different histones or even different lysines in the same histone tail, For example - PCAF targets H3 and H4; SAGA targets H3 and H2B , both modification creates bromodomain.
• HDAC (Histone deactyltransferase) - HDAC removes the modification created by HAT; removes acetyl gropus from the histones.
• DNMT (DNA methyl transferase) - This enzymes add methyl groups to the histones, targets specific lysine or arginine on specific histone. This modifications creates chromodomain. Methylation of H3 at 9th lysine results in gene silencing whereas methylation on H3 at 4th lysine turns ON the transcription.

b)

• The ATP dependent nucleosome remodelling complex, Swi/Snf, influences the stability of the histone octamer-DNA interaction and inturn this remodelling of nucleosome allows increased access to the DNA
• Acetylation reduces the overall positive charge of the histone tails which in turn reduces the affinity of the tails for the negatively charged DNA. This makes RNA polymerase and transcription factors easier to access the promoter region
• HDAC results in compact chromatin that suppresses transcription
• Unmodified or methylated histone tails associate tightly with nucleosomal DNA resulting in the gene silencing

c)

• HAT are associated with the regions of the chromosomes that are transcriptionally active
• HDAT nucleosomes are associated with transcriptionally repressed chromatin
• DNMT catalyzes methylation of different parts of N-terminal tail, is associated with both repressed and active chromatin, depending on the particular amino acid that is mofified in the histone tail