Control of gene expression

Question 1

TRANSCRIPTION REGULATORS

Question 2

CO-ACTIVATORS

Question 3

CO-REPRESSORS

Question 4

THE MEDIATOR COMPLEX

Answer 4

The mediator complex serves a crucial function in gene regulation, forming a link between
gene-specific transcription factors and RNA polymerase II.
The gene regulatory proteins interact with
the mediator complex and may recruit
additional regulatory proteins.

Question 5

CHROMATIN AND GENE REGULATION

Question 6

TRANSCRIPTION CIRCUITS AND GENOMIC SWITCHES

Question 7

POSITIVE CONTROL

Answer 7

A proliferating cell must maintain its identity through subsequent cell division, it has some form of cell
memory, and its pattern of gene expression must be transferred to its daughter cells.
* Positive feedback loops may be used to establish and maintain a heritable pattern of gene expression.

Question 8

NEGATIVE CONTROL

Answer 8

Positive and negative regulatory feedback loops are common in all cells and can be combined in different
gene regulatory circuits.
* A negative feedback loop is often used to keep the concentration of a protein within a defined range.
Low levels (lack of repressor) → higher expression
High levels (active repressor) → lower expression

Question 9

COMBINATIONAL CONTROLS

Question 10

EPIGENETIC MECHANISM

Question 11

CELL MEMORY

Question 12

POST TRANSCRIPTIONAL REGULATION

Question 13

RNA INTERFERENCE

Answer 13

RNAi is induced by a cell detecting double stranded RNA (dsRNA).
* The dsRNA are processed into small 20-30 bp single stranded RNAs, which binds to specific proteins
(Argonaute and PIWI ) and act as guide RNAs to identify other complementary RNAs in the cell.
The three major classes:
microRNAs (miRNAs), small interfering RNAs (siRNAs) and piwi interacting RNAs (piRNAs)

Question 14

CRISPR SYSTEM

Answer 14

CRISPR: Clustered Regularly Interspaced Short Palindromic Repeats
* CRISPR-type clustered repeats were first reported in E. coli (1987), but their functions are only recently
uncovered.
* During infection short fragments of viral DNA are integrated into the bacterial genome.
A later infection with the same / similar bacteriophage will be detected by the CRISPR/Cas9
“surveillance system” and phage DNA degraded.
* A protein complex including the TracrRNA helps processing pre-CRISPR RNA. RNA from the spacers
(of viral origin) are bound to a nuclease (Cas9), which uses the crRNA to seek and destroy viral DNA.

Question 15

ENHANCERS

Question 16

INSULATORS

Answer 16

Insulators restrict the range of activators / repressors, so they do not affect nearby genes.
* Insulators function by forming loops of chromatin, bringing genes and control regions in close proximity.

Question 17

DNA METYLATION

Answer 17

Methylated DNA is recognized and bound by
specific proteins. This may block other gene
regulatory proteins access to DNA.
* A repressive form of chromatin may be established
through histone modifications, by DNA methylases
and proteins binding methylated DNA.
* A histone modifying enzyme is recruited by a gene
regulatory protein (a repressor).
* The modified histone is “read” by a code reader
protein and histone modification is transferred to
neighboring histone by the histone reader-writer
proteins.
* A DNA methylase is recruited and cytosines are
methylated.
* DNA methyl-binding proteins bind methylated
cytosines and cause gene repression.

Question 18

HISTONE MODIFICATIONS

Answer 18

Histone modifications provide favorable interactions for a number of proteins, (chromatin
remodeling complexes, reader-writer complexes).
* Modifications can be rapidly reversed (genes that needs to be quickly turned on or off), or
modifications can persist and serve as a short and long-term cellular memory.
* Histone modifications can even be transferred to daughter cells after cell division.

Question 19

LONG NON-CODING RNA

Answer 19

RNAs longer than 200 bp that apparently do not code for any protein.
* Many are transcribed by RNA pol II and have 5’ cap and poly A tails.
* At least 16000 lncRNA have been discovered in humans (GENCODE).
* lncRNAs may function as scaffold RNAs providing interaction between proteins and coordinate their
function or may have guide functions.

Question 20

XIST

Answer 20

Long noncoding RNAs (lncRNAs) such as Xist are involved in X-chromosome inactivation.
X-chromosome inactivation spreads from the X-inactivation center (XIC) where a long noncoding RNA
(Xist) is expressed.
Only one of the X-chromosomes have Xist
expression.
* The Xist lncRNA diffuses along the Xchromosome and recruit proteins that
induced chromatin modifications and DNA
methylation, resulting in gene inactivation.

Question 21

RNA DEGRADATION

Answer 21

Decapping and degradation of mRNA takes place at specific locations in the cytosol, so called
Processing bodies (P-bodies).
* If they are not degraded they can be transported over to so called stress granules and be
”restored” for later use.
There is a competition between
mRNA translation and mRNA
degradation.
* If a mRNA is translated with high
efficiency, it is less prone to
degradation.

Question 22

ALTERNATIVE mRNA SPLICING

Answer 22

Alternative splicing is common in human genes (90 % of the genes).
* Alternative splicing increases the protein complexity.
* Some genes containing many exons can have a large number of splice
variants.
* In many cases alternative RNA splicing is regulated and various splice
forms are produced in different cell types.
In back splicing a single exon is removed as a circular RNA molecule.
The biological functions of circular RNAs (circRNAs) are still not fully
understood but they may regulate function of microRNAs (miRNAs).
Negative control of RNA splicing:
* A repressor prevent the splicing machinery access to the splice site and obstruct mRNA splicing. May
instead result in the use of a “cryptic” splice site.
Positive control of RNA splicing:
* A regulatory protein (splicing enhancer) is required for mRNA splicing. The enhancer binding site can be
located far from the splice site.

Question 23

RNA EDITING

Answer 23

RNA editing alter the nucleotide sequence of RNA transcripts and can change their coding sequence,
produce stop codons or affect mRNA splicing.
* In animals the most common editing is deamination of adenine to produce inosine (A-to-I editing) and
deamination of cytosine to produce uracil (C-to-U editing).
* A-to-I editing is most common in humans and occur in approx. 1000 genes
The ADAR enzymes (adenosine
deaminase acting on RNA) carry out
the A-to-I editing.
A double stranded RNA structure
produced by the mRNA is recognized
by ADAR enzyme and defines the site
for RNA editing.
Editing take place in the nucleus before
the pre-mRNA are fully processed.

Question 24

siRNA

Answer 24

The presence of dsRNA in the cell (ex. from a virus)
triggers RNAi by recruiting a protein complex
containing Dicer.
* This protein complex cleaves the dsRNA into ~23
bp fragments called small interfering RNAs (siRNA).
* Binding of the siRNA to the RISC complex
results in degradation of one of the RNA
strands and enable the 5’ end of the RNA to
bind complementary mRNA sequences →
degradation of mRNA.
An alternatively RNA interference response is directed by the RNAinduced transcriptional silencing (RITS) complex.

Question 25

miRNA

Answer 25

Non-coding RNAs transcribed by the cell, which
regulate protein coding genes.
* One class of short non-coding RNAs (21-23 bp)
is the miRNAs.
* miRNA precursor are synthesized by RNA pol II
and polyadenylated.
* They form a distinct stem and loop structure
which is cleaved.
* In cytosol the miRNA cleaved by the Dicer
enzyme and is bound to the RISC complex which
includes the Argonaute protein.
* In the RISC complex one of the RNA strands are
degraded leaving the 5’ end of other strand to
bind complementary mRNAs.
* This induces the Argonaute protein to cleave the
mRNA and promote mRNA degradation.

Question 26

piwiRNA

Answer 26

Piwi-interacting RNAs (piRNAs) are particularly expressed in the germ line of animals.
* The pre-transcripts of piRNA are processed by nucleases that cleave them into 24–32 nt in
length.
* The Piwi proteins are structurally related to Argonaut proteins and bind piRNA.
* They can degrade specific mRNA or be involved in epigenetic regulation and silencing of genes.
* The piRNA are clustered in the genome and has an important function in keeping
transposable elements (transposons) in check, primarily during the development of germ
cells which undergo cytosine demethylation that may lead to transcriptional activation of the
transposons.

Question 27

RISC COMPLEX