CMB2001/L10 Post-transcriptional Control IIb

Question 1

Using casein mRNA, explain how mRNA levels are regulated.

Answer 1

Expressed in mammary gland
mRNA increases 70-fold on stimulation by prolactin
Transcription increases 2-fold as half-life increases 40-fold
PolyA tail length increased
3’ UTR of RNA binds proteins which aid in stabilisation

Question 2

Give 3 reasons for mRNA degradation.

Answer 2

Damaged mRNA
Incorrectly transcribed/processed mRNA
Control gene expression

Question 3

What must occur to mRNA before exonucleases can gain access?

Answer 3

Closed loop must be broken

Question 4

Describe deadenylation-dependent decay.

Answer 4

Decapping of mRNA by DCP1&2 and 5’-3’ degradation by exonucleases
PolyA tail removal by CCR4-NOT or PARN by removing adenine residues (then exosome complex degrades 3’-5’)

Question 5

Give 2 decapping enzymes.

Answer 5

DCP1
DCP1

Question 6

Give 2 endonucleases.

Answer 6

Argonaute
Swt1
Smg6

Question 7

Give a deadenylase enzyme.

Answer 7

CCR/Not complex

Question 8

Which enzyme degrade mRNA:
a) 5’-3’
b) 3’-5’?

Answer 8

a) XRN1
b) the exosome

Question 9

Give 2 nucleases in the exosome.

Answer 9

RRP6
RRP44

Question 10

Give another use of the exosome.

Answer 10

RNA turnover and processing

Question 11

Give 2 other functions of XRN1.

Answer 11

Involved in RNA turnover and processing
Involved in transcription termination
Functions after decapping of mRNA

Question 12

Describe nonsense mediated decay (NMD).

Answer 12

Mistakes in RNA are detected and mRNA targeted for degradation

Question 13

Give 3 ways in which premature stop codons can occur in mRNA.

Answer 13

Errors in:
Transcription
Splicing
Editing
Polyadenylation
Mutations

Question 14

Describe nonsense-mediated decay as a surveillence mechanism.

Answer 14

Targets mRNA with premature stop codons (nonsense)
Recognises presence of stop codons and tags mRNA for degradation
Stops production of truncated, potentially harmful proteins

Question 15

Describe the role of the exon junction complex (EJC).

Answer 15

Deposited near splice junctions during mRNA splicing
Marks spliced mRNAs
If ribosome encounters premature stop codon and EJC remains downstream, indicated mRNA is defective

Question 16

What do these abbreviations mean:
a) RNAi
b) siRNA
c) miRNA
d) RISC?

Answer 16

a) RNA interference
b) small inhibitory RNA
c) micro RNA
d) RNA-induced silencing complex

Question 17

Describe siRNA.

Answer 17

21-23 nucleotide RNAs
Perfect complementary to target RNA
Thought to be mainly viral defence mechanism
Leads to degradation of target RNA

Question 18

Describe miRNA.

Answer 18

21-23 nucleotide RNAs
Imperfect complementary to target RNA
Key gene regulatory mechanism in the cell
Leads to block in translation

Question 19

Describe siRNA production.

Answer 19

dsRNA precursor
Dicer enzyme recognises and processes long dsDNA into fragments
Incorporation into RISC
Target recognition and cleavage

Question 20

Describe miRNA production.

Answer 20

Primary miRNA (pri-miRNA)
Drosha enzyme with cofactor DGCR8 processes pri-mRNA into shorter precursor
Export to cytoplasm
Dicer enzyme processes into mature miRNA duplexes
One strand incorporation into RISC - other is degraded
Target recognition and regulation

Question 21

What happens to UTRs during embryonic development?

Answer 21

Get longer

Question 22

How does the length of mRNA differ in proliferating cells?

Answer 22

They’re shorter

Question 23

Give 3 uses of siRNA in research.

Answer 23

Gene knockdown
Functional genomics
Disease modelling
Target validation
Therapeutic applications
Pathway analysis
High-throughput screening