RNA modifications/degradation

Question 1

What do we have at the 3’ end?

Answer 1

The polyadenylation signal

Question 2

What is the first modification of RNA?

Answer 2

the 5’ cap

Question 3

What are the functions of the polyadenylation signal?

Answer 3

1. It will determine the cleavage of new synthesised RNA at some point 20-30 nucleotides downstream of the message AAUAAA sequence
2. A poly-A tail will be synthesised at the end of the RNA

Question 4

What do RNA modifications ensure?

Answer 4

Ensure that RNA is to be protected and stable in the cytosol

Question 5

Why does the RNA get a circular conformation?

Answer 5

After translation stops, the ribosome disassembles and the point where it disassembles is closer to the point of translation start rather then the linear conformation

Question 6

What is the 5’ cap?

Answer 6

7-methylguanosine: cannot be degraded by exonucleases

Question 7

What do mechanisms of RNA degradation imply?

Answer 7

Deadenylation or decapping

Question 8

Which protein complexes are involved in de-adenylation?

Answer 8

PARN digests the polyA tail

Question 9

Which enzymes are involved in decapping at the 5’?

Answer 9

DCP2

Question 10

Exonucleases

Answer 10

exert degradation either in 5’ to 3’ direction through Xrn1 or from the 3’ to the 5’ through exosomes

Question 11

Endonucleases

Answer 11

Cleave the mRNA in the middle: generate 2 molecules of RNA one without the polyA tail and one without a 5’ cap

Question 12

Digestion of the polyA tail

Answer 12

1. Start the degradation from the 3’ end
2. Binding of a protein complex called LSM127
3. Leads to the decapping of the proteins on the 5’ end
4. leads to 5’ to 3’ degradation

Question 13

When does non-stop decay take place?

Answer 13

When you have a mutation which results in the removal of the physiological stop codon

Question 14

What happens in non-stop deacy?

Answer 14

1. When the ribosome gets to the very end of the RNA molecule it is stopped
2. This causes the binding of the Ski7 protein
3. Ski7 recruits the exosomes which start the 3’ to 5’ degradation
   When Ski7 is absent we have the loss of the polyA binding proteins and this allows decapping at the 5’ end by Xmr1

Question 15

What is no-go decay?

Answer 15

A mutation that leads to the formation of a strong secondary structure that causes the stopping of the ribosome so it cannot continue with translation

Question 16

What happens in no-go decay?

Answer 16

The stalled ribosome will recruit 2 proteins: Dom34 and Hbs1 which catalyse the endonucleolytic cleavage of the mRNA–> binding of either exosomes of Xrn1

Question 17

Non-sense mediated decay

Answer 17

Triggered by the presence of a premature stop codon somewhere before the physiological one in the mRNA

Question 18

What are the constituents of the EJC?

Answer 18

Upf2 and Upf3 where Upf2 is capable of binding to Upf1

Question 19

What happens in NMD normal mRNA?

Answer 19

1. The ribosome is bound at the top and starts translating towards the EJC which has bounded the Upf2 and Upf3
2. The ribosome in the pioneering round dissociates the EJC and gets to the physiological stop codon

Question 20

What happens when we have a premature stop codon?

Answer 20

1. When we have a premature stop codon, the ribosome gets to it.
2. There is an EJC downstream, so we have the binding of a release factor which recruits Upf1
3. The Upf1 recruits the kinase SMG-1 which together with release factors form a transient complex called SURF
4. The SURF complex interacts with the EJC downstream and Upf1 interacts with UPf2
5. SMG-1 phosphorylates Upf1 which triggers mRNA degradation
6. The phosphorylation leads to the dissociation of the ribosome and of release factors
7. What remains is parts of the SURF complex and the recruitment of 2 SMG proteins: SMG-5 and SMG-7+ phosphatase PP2A
8. This leads to dephosphorylation of Upf1 and decapping which is triggered by SMG-2

Question 21

SURF complex

Answer 21

SMG-1, Upf1, eRF complex, Upf2, Upf3

Question 22

Where are the proteins involved in NMD concentrated in?

Answer 22

processing bodies

Question 23

What happens if there is a premature stop codon in a monoexonic gene?

Answer 23

These genes skip NMD as they do not have the signature protein complex required to recognise premature stop codons

NMD resistant

Question 24

What do monoexonic genes code for?

Answer 24

Histone proteins
Ribonucleases
Heat shock proteins HSP

Question 25

How can a premature stop codon be skipped?

Answer 25

If we have a readthrough mechanism

Question 26

What is the readthrough mechanism?

Answer 26

Induced by a treatment of aminoglycosidic antibiotics that inserts a near-cognate tRNA [tRNA complementary for 2 nucleotides of the recognised stop codon] which will compete with release factors

Question 27

What are aminoglycosidic antibiotics?

Answer 27

Capable of inducing the insertion of a near-cognate tRNA which does not match the anticodon–> leads to the skipping of the premature stop codon

Question 28

What is the effect of aminoglycosidic antibiotics?

Answer 28

The antibiotic interacts with the ribosome and catalyses the polypeptide chain

Question 29

Examples of aminoglycosidic antibiotics

Answer 29

Gentamycin
Akamycin

Question 30

What have gentamycin and akamycin been used for?

Answer 30

Treatment for CFTR gene and dystrophin gene

Question 31

What is a downside of aminoglycosidic antibiotics?

Answer 31

toxicity

Question 32

What can be used instead of aminoglycosidic antibiotics?

Answer 32

PTC124 molecule

Question 33

Describe PTC124 molecule

Answer 33

Able to skip the effect of the premature stop codon and leads to the production of the protein by insertion of a random aa

Question 34

Features of PTC124 molecule

Answer 34

1. Non aminoglycoside
2. no antibacterial activity