Regulation of gene expression

Question 1

7 steps of the overall regulation of gene expression

Answer 1

1. transcription initiation (DNA is converts into primary RNA transcripts)
2. Posttranscriptional processing (primary RNA transcript into mRNA)
3. RNA stability
4. translational regulation (mRNA into a protein)
5. protein modification
6. protein transport
7. protein degradation
   * * regulation at transcription level is the best understood process!!!

Question 2

definition of promoter and its access is restricted by what

Answer 2

DNA sequence at which RNA polymerase may bind, leading to the initiation of the transcription
- its access is restricted by chromatin

Question 3

transcription and translation are separated by what

Answer 3

by the nucleus

Question 4

where does the RNA polymerase and the regulatory proteins bind + what these proteins do

Answer 4

RNA polymerase binds to DNA at promoter

regulatory proteins bind to promoter to regulate RNA pol2, TF, activators and repressors (operators)

Question 5

4 molecular signal for the regulation

Answer 5

hormone
covalent modification
allosteric regulator
interacting protein

Question 6

what are the 4 scenarios of regulation (2 negative and 2 positive regulation)

Answer 6

negative regulation:
1. molecular signal causes dissociation of repressor (from the operator, the zone where the repressor binds) from DNA, inducing transcription
2. molecular signal causes binding of repressor to DNA, inhibiting transcription
positive regulation (with activators)
3. molecular signal causes dissociation of activator from DNA, inhibiting transcription
4. molecular signal causes binding of activator to DNA, inducing transcription

Question 7

what are the regulatory sequences in the promoter

Answer 7

1. activator binding site

2. repressor binding site (operator)

Question 8

how the regulatory proteins bind to DNA

Answer 8

regulatory proteins have DNA binding domains/motifs

interaction specificity depends on hydrogen-bonds donors and acceptors in bases (DNA) and AA (proteins)

Question 9

5 most commons AA involved in the binding of regulatory proteins to DNA

Answer 9

Asn, Arg, Glu, Gln, Lys

Question 10

3 common types of DNA binding motifs

Answer 10

1. zinc finger domain
2. helix-turn-helix domain
3. leucine zipper

Question 11

the composition of the helix-turn-helix domain and one example

Answer 11

20 AA long with 2 a segments
one of the a segments is called the recognition helix because it usually contains many of the amino acids that interacts with the DNA in a sequence-specific pathway
** so even if there is 2 a segments, only one has the function of DNA binder
- example : lac repressor

Question 12

composition of zinc finger domains

Answer 12

• 30 AA form an elongated loop held together at the base by a single Zn2+ ion, which is coordinated to 4 of the residues (4 Cys, or 2 Cys and 2 His)
• The zinc does not itself interact with the DNA; rather, the coordination of zinc with the amino acid residues stabilizes this small structural motif (+ hydrophobic interactions are there to stabilize)

Question 13

characteristics of the leucine zipper

Answer 13

• Leu occurs at every 7th position
• partially interacts with DNA (Lys/Arg) (Regulatory proteins with leucine zippers often have a separate DNA-binding domain with a high concentration of basic (Lys or Arg) residues that can interact with the negatively charged phosphate of the DNA backbone)

Question 14

what do regulatory have (2)

Answer 14

• protein-interaction domain (so can interact with other proteins to form complexe and do the transcription
• DNA binding motifs

Question 15

chromatin = ?

Answer 15

DNA + histones (H1,H2A,H2B,H3,H4)

Question 16

whar are the variants of histone

Answer 16

H3.3 and H2AZ

Question 17

which enzymes deplete H1 and replace histone variants

Answer 17

SWI/SNF
** These enzymes remodel chromatin so that nucleosomes become more irregularly spaced and they also stimulate transcription factor binding

Question 18

histones are covalently modified (5 different modifications + AA implicated)

Answer 18

1. methylation (lysine and arginine)
2. phosphorylation (serine and threonine)
3. acetylation (lysine)
4. ubiquitination (lysine)
5. sumoylation (lysine)

Question 19

where the methylation of DNA occurs

Answer 19

at CpG sequence

• • It plays a gender role in transcription (methylation of the DNA itself, it occurs in CpG sequences (in the nucleotides pair where C is followed by a G and not the other way). Of these nucleotides, it is the C (cytosine) that is methylated
• • the methylation inhibit the access to the promoter region

Question 20

general steps of pre-initiation complex

Answer 20

1. activator bind to the enhancer
2. histone/nucleosome modification at promoter
3. TATA binding protein (TBP) and TF2B bind to promoter
4. basal TF and RNA pol2 bind at transcription start site (TSS)
5. TF2H stimulates Pol2 to transcribe

Question 21

what are the three domains of nuclear receptors (that bind to HREs, which are specific sequences of DNA)

Answer 21

1. transcription activation (variable in lenght and sequence)
2. DNA binding (66-68 residues, highly conserved)
3. hormone binding (variable sequence and lenght)

Question 22

description of the type 1 nuclear receptors

Answer 22

1. they are found in the cytoplasm in an inactive form link to a protein Hsp70
2. the hormone comes in the cytoplasm and link to the receptro, which dissociates the Hsp70
3. the nuclear will dimerize (link with another)
4. it will diffuse in the nucleous where it will bind to HREs and act as transcription activator

Question 23

description of type 2 nuclear receptors

Answer 23

1. they are found in the nucleus in an inactive form and bound to a crorepressor and to HREs
2. the hormone migrates through the cytoplasm and diffuses accroos the nucleus membrane and its binding with the receptor (binds to a heterodimer) will change the conformation so it will lead to the dissociation of the corepressor
3. nuclear receptor acts as a transcription activator

Question 24

why does one gene can lead to different protein (what is the phenomenon + give an example)

Answer 24

alternative splicing at transcript level:
95% of with multiple exons undergo alternative splicing
- we have the primary transcript (with many exons and introns) and before it goes to mRNA for the translation, there are some transformations where some exons are retained and others are not and where the introns are taking out
so at the end, the mRNA can be different even if it starts from one gene
ex: by going in the thyroid, the calcitonin is the synthesize protein (carrier hormone)
by going in the brain, the CRGP is synthesizes
( the difference is tha the exon 4 has not been conserved in the brain)

Question 25

where is the translation process and what regulates it

Answer 25

after the transcription and the post-transcription, it goes in the cytoplasm
- there is the binding of translational repressors to the to specific site in the 3prime untranslated region of the mRNA. These proteins interact with eukaryotic initiation factors or with the ribosome to prevent or slow translation

Question 26

what can inhibit the translation and how

Answer 26

micro-RNAs (miRNAs)
it can prevent the translation of mRNA by cleaving it (via endonuclease Drosha or Dicer) or by blocking it
** they bind to target sequences
** it interacts with mRNAs often in the 3primeUTRm resulting in either RNA degradation or translation inhibition