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Regulation of Gene Expression
Summary Slides in Question Form
1. What are activator binding sites? Where are they located? Why?
These are usually upstream from the RNA pol binding site. It's upstream so that it can activate
the RNA pol site...if it was downstream, the RNA pol would already have been activated.
2. What is the difference between negative and positive regulation?
Negative regulation is done by a repressor to impede the gene; positive regulation is done by an
activator which enhances the pol/promoter interaction. A corepressor enhances the binding of a
repressor, so that it repressors further. An inducer of a repressor will lead to more gene regulation,
because it blocks the repressing action. A co-activator, likewise, helps an activator work better; a
repressor reduces the inducing work of the activator, so it leads to less gene expression.
3. How does the Tryptophan Operon work?
Operons are only in bactera. It's a cluster of genes in a polycistronic mRNA. An operon
includes the promoter region which has binding sites for an activator, a repressor, and RNA pol.
The Tryp Operon is a classic example. The promoter is a 15 bp region recognized by a
repressor protein. The repressor only works when it has two Trp molecules bound to it. So, when there
is Trp in the cell, the gene to make more Trp is turned off. But, when there is less Trp in the cell, the
repressor is NOT bound, so the gene to make Trp gets turned ON.....it makes Trp, then those same Trp
that it made go back to bind to the Operon, which shuts OFF the gene, since it has what it needs. This
is negative control.
4. How does the Lactose Operon work? What is are the signal proteins, and what do the effect?
In what situations is the Lac operon under positive/negative control?
The Lac Operon is another example of negative control, where the gene is turned off until there
is a signal that its product is needed. E Coli prefers to use glucose, but when it runs out, it has a
mechanism to use lactose, instead. Both glucose and lactose are involved in the regulation. The lac
operon can only work when it's activated by CRP/CAP, which can only bind when it has cAMP.
Glucose lowers cAMP. When there's less glucose, there's more cAMP. Lactose, through allolactose,
causes dissocation of the repressor from the operator. So, with glucose and lactose, gene is off. With
glucose, but no lactose, gene is off. With neither glucose or lactose, gene is off. With no glucose, but
only lactose, there is allolactose, plus cAMP, gene turns on.
5. What is the role of molecular signals with repressors/activators? Where do repressors/activators
bind in relation to the RNA pol binding site?
Repressors work with signalling from other molecular signals, and they bind to an operator site
near the binding site for RNA pol.
6. How does attenuation work? What does the attenuator region consist of?
Works as an extra brake function to stop transcription that started accidently. The region
consists of an ORF (open reading frame) for the leader peptide and three downstream regions that can
make different hairpin options.
7. What happens when there is enough of the amino acid that an operon codes for? When there is
not enough of the amino acid the operon codes for?
So with the Trp attenuator, if the repressor lets RNA pol start going accidentally, two things
could happen: if there is Trp and the cell doesn't need more, then the RNA pol goes through sequence 1
quickly, which blocks sequence 2....this lets 3 and 4 form a hairpin that blocks further transcription. If
there is low Trp in the cell, then the reading of sequence 1 is slow, which allows 2 and 3 to form a
hairpin, which is a little bump that the RNA pol can get over to continue transcription. 2 and 3
happens naturally, but 3 and 4 form the attenuating hairpin.
8. What does “negatively mediated” mean?
Actually, “negatively regulated” is mediated by the one of the proteins that the gene/operon codes for.
9. What's missing in DNase hypersensitive sites?
Regions in chromosomes that are devoid of nucleosomes and allow for the binding of Tfs. About 100200 bp, within 100 bp upstream from the iniation site, but could be somewhere else (if DNA is folding
over). These are tissue specific and can be used to figure out tumor origin, etc.
10. What is hemoglobin switching, and what happens between embryonic, fetal, and adult stages?
This is developmentally regulated and is associated with Dnase-hypersensitive sites. Embro
starts out with zeta/epsilon Hgs. The, at 5-6 weeks, the fetal liver switches to gamma Beta-globin. Just
before birth, both zeta and gamma globins are replaced by alpha and Beta Hg from the bone marrow.
11. How can promoter elements be classified? Who are the two types devoted to?
The can be classified as proximal or distal promoters.....proximal for generic Tfs and distal for
gene-specific Tfs.
12. What are co-activators involved in?
Co-activators are either involved in binding of transactivators to the RNA pol II complex and
TAFs or in chromatin remodeling.
13. Where are nuclear hormone receptors translocated to, and when? How do they bind to DNA as,
and where?
Translocated in the nucleus after binding a steroid hormone. They bind to DNA as dimers to a
repeated DNA sequence. Nuclear hormone receptors have 2 conserved domains: 2 zinc fingers/DNA
binding domain and a ligand binding domain. DNA binding sites are HREs.
***Glucocorticoid receptor normally abound by HSP 90, inactive. But, dimerisation activates
genes which have HRE element through chromatin remodelling coactivators. Ligand inducible Tfs.
14. What activities to co-activators and co-repressors have?
Coactivators have HAT (histone acetyl transferase) ativity; co-repressors have HDAC (histone
deacetylase) Histone acetylation prevents chromatin from folding compactly, which looses it up for
gene expression. So, HDAC would close it back up, deacetylation.
***CBP is a HAT.....deficiency of CBP results in Rubinstein-Taybe Syndrom.....big thumbs, MR, facial
growth
15. How does Tamoxifen work?
Estrogen analog, binds in the pocket normally used by estrogen. This distorts the structure of
the estrogen receptor, which blocks the binding of the coactivator, aka, it blocks the recruitment of
HATs.
16. How do non-steroid hormones regulate?
They regulate gene expression by intracellular signaling through phophorylation of the protein,
followed by chromatin remodeling. Ex: Epi activates GPCR, which increases cAMP, PKA....PKA
phosphorylates the CREB in the nucleus, which binds to CBP.....this causes histone acetylation and
remodeling, opening up the DNA for gene expression. Thanks, epi.
17. How many genes do transcription factors regulate? What do they bind to DNA through and
where?
One TF may regulate many genes. They bind to DNA through glutamine, asparagine, arginine,
lysine, and glutamine (all the long side chains) by H-boding and hydrophobic interactions in the major
DNA groove.
18. What are helix-turn-helix and Zinc finger domains, , and what are they involved in?
Bacteria proteins interact with DNA with helix-turn-helix motifs.....this interacts with DNA in
the recognition helix. There is one of these in the lac repressor. HtH and Zinc finger proteins are
infolved in TF-DNA interactions.....helps the TF bind to the DNA. A 60 aa HtH is called a
homeodomain, and its gene is a homeobox. HOX genes are super important for embryonic
development.....mutations in HOXD13 => fused fingers.
19. What do HMG-box protein domains do?
High Mobility Group motifs bend DNA and stabilize it for mediators.....these are present in Tfs
that bend DNA. SRY, the male-ness gene, bends with HMG.
20. What do leucine zipper and helix-loop-helix domains do?
These are domains involved in interaction between different Tfs.Fos and Jun protiens contain bZIP
domains. Helix-loop-helix region also helps DNA binding....MyoD, the TF for muscle, has this.
21. Where are CpG dinucleotides methylated, and why?
CpG are methylated at the C residue....methylated DNA is associated with gene silencing;
demethylation activates DNA.
22. MeCP binds to _________. What does it do?
Binds to methylated C in the CpG dinucleotides, then assists in deacetylation/then methylation
of core histone proteins.
23. In what type of cells does regulation of translation occur in primarily, and what does it involve?
Eukaryotic cells: involves either phosphorylaton or Ifs or interaction with hairpins in the UTRs
of mRNA. ElF2 phosphorylation is necessary for globin synthesis; Heme can regulate this depending
on iron levels. Yes, iron? Heme present, kinase inhibited by elF2. No, iron? No heme, elF2 is
phosphylated, degraded.
24. What are iron-responsive elements, and where are they located?
Ferritin....stores iron when there is too much. Transferrin receptor takes up iron when there's
not enough, or to save it for later. So, if there's not enough iron, you dont want Ferritin to get it, you
want transferrin to get it so you can use it. If there's too much iron, you don't want transferrin to get it
and use it (toxic), you want Ferritin to get it and store it safely. Both Ferritin and Transferrin mRNA
have an IRE to regulate it. Ferritin has IRE in the 5'.....excess iron causes the binding protein of the
IRE to let go from where it was binding to block new ferritin from being made. More iron? 5' IRE of
ferritin removes binding protein, so more ferritin is translated. In low iron, binding of the 3'IRE
protein stabalizes the transferring receptor mRNA, so more transferrin is made.
25. What happens with ferritin receptor proteins at low and high iron concentrations?
High iron? Up-reg ferritin, down-reg transferrin-R. Low iron? Upreg transferrin, down reg ferritin.