Virology: Chapter 6

Question 1

What is the structure of the poliovirus

Answer 1

naked, 30 nm, icosahedral spherical, densely packed to 60 promotes

Question 2

what is each promoter made of

Answer 2

VP0, VP1, VP3

Question 3

how many capsomeres does the virus have

Answer 3

20

Question 4

what is the difference between a mature and immature virus in the context of proteins

Answer 4

mature: VP0 is cleaved to VP2 (exterior) and VP4 (interior) but this happens later in the replication cycle

Question 5

what is the use of sodium and potassium inside the capsid

Answer 5

to fight the (-) charge on the phosphate group of the RNA

Question 6

what is the genome of poliovirus

Answer 6

(+) sense single stranded RNA

Question 7

what is meant by a (+) RNA genome?

Answer 7

it has a coding sequence and is used as a template to transcribe viral mRNA

Question 8

where does poliovirus replicate and express its genome in the host cell?

Answer 8

cytoplasm

Question 9

is the genome of the poliovirus capped or uncapped

Answer 9

uncapped

Question 10

and what is the protein that is covalently attached to 5’ end

Answer 10

VPg protein

Question 11

which UTR is longer, 5’ or 3’

Answer 11

5’ is has a longer UTR than the 3’

Question 12

what is the 5’ UTR for?

Answer 12

it’s the structural gene
- for translation and genome packaging into capsids
(attract IRES and translate for capsid proteins)

Question 13

what is the 3’ UTR for

Answer 13

• non-structural gene
• synthesis of the (-) strand for the replication of genome

Question 14

what is the purpose of (-) sense RNA strand in poliovirus genome replication?

Answer 14

it’s used a template to create more (+) RNA to be the genome of future viruses and to be mRNA for further translation

Question 15

What does it mean for the RNA genome to be infectious

Answer 15

it means if its injected in the host cell, progeny will always be produced

Question 16

what is the receptor protein of poliovirus

Answer 16

CD155 (PVR)

Question 17

where is CD155 found

Answer 17

macrophages, monocytes, CNS neurons
- immune cells

Question 18

Canyon VP1 binds to CD155 triggering conformational change (pH dependent) of the capsid, resulting in

Answer 18

N-terminus of VP1 and VP4 exposed to surface, and VP1 insert into plasma membrane to form a pore, viral genome enters cytoplasm

Question 19

after that, VPg is cleaved off by protease, what happens after that?

Answer 19

IRES assembles, acting as RBS

Question 20

where does IRES and ribosomes assemble

Answer 20

5’ UTR, because the ribosomes will read from 5’ to 3’

Question 21

translation of (+) RNA genome immediately happens, and produces a polyprotein, how many ORFs does the (+) RNA have?

Answer 21

1

Question 22

as it synthesizes the polyprotein, it undergoes folding and becomes activated, what does it do next

Answer 22

cleaves to form 20 different proteins

Question 23

what are the major groups of those proteins

Answer 23

structural genes: VP0, VP1, VP3
non-structural genes: RDRP, VPg, Proteases

Question 24

to eliminate competition of ribosomes with host cell mRNA, what does protease do?

Answer 24

damages the CBC of host cell mRNA (elF4G) so the ribosomes can bind to the viral RNA instead

Question 25

after that, forms a replication compartment, what is it made out of?

Answer 25

RDRP, VPg, (+) strand RNA

Question 26

why is the replication compartment membrane-bound?

Answer 26

to avoid detection and being degraded by cellular RNAses (protect)

Question 27

for replication, poliovirus RDRP needs a primer, what is that primer and what does it do

Answer 27

VPg, it allows polymerase to attach nucleotides to 3' OH VPg --> VPG--U--U (add 2 uracils and 3'OH == primer!!)

Question 28

Then, RDRP reads (+) strand and adds complementary base to primer, producing.... that acts as....

Answer 28

(-) ds RNA that acts as a template to synthesize new (+) ss RNA

Question 29

After that, there are two scenarios that happen

Answer 29

Vpg is cleaved off, so newly made (+) RNA is translated to make more capsid proteins for the progeny virus VPg stays on, so the (+) RNA is used for the new genome for the progeny virus

Question 30

The strutural genes that were cleaved from the polyprotein during the earlier stage, VP0, VP1, and VP3 play a role in assembly, which is...

Answer 30

form the promoter --> pentamer (14S) --> procapsid procapsid is capsid without genome

Question 31

So we want the genome to enter the procapsid, how do we ensure (+) ss RNA is inserted in, and NOT (-) ds-RNA

Answer 31

the (+) ss RNA contains a PAC site that interacts with capsid proteins

Question 32

Why is it vital that the (+)-sense and not the (–)-sense RNA be packaged into the pro-capsids? What allows for the proper packaging of the (+) sense RNA?

Answer 32

(+) sense RNA cannot have RDRP (to save space) and a PAC site that interacts with capsid proteins

Question 33

Maturation happens, which is indicated by

Answer 33

VP0 cleaved to VP2 and VP4

Question 34

what do these proteins do (VP2 and VP4)

Answer 34

seal up the capsid and protect RNA from degradation

Question 35

The cell is released by lyses, what is a protein that is involved in it, and what does it do?

Answer 35

Viroporins, form pores to disrupt cell's membrane to release poliovirus

Question 36

Would poliovirus be able to use the host cell RNA polymerase to replicate and transcribe its genome? Explain

Answer 36

No, because host cell polymerase does not read RNA as a template, that's why is it's a (-) sense, it must pre-package RDRP in the capsid, but since polio is (+) sense, it makes RDRP on the spot.

Question 37

The poliovirus mRNA has no 5’-methylated cap. How do the ribosomes assemble onto the RNA for translation?

Answer 37

they rely on a secondary structure called IRES, that interacts with some cellular proteins to assembly onto a ribosome (attract ribosomes)

Question 38

Why does the poliovirus utilize a polyprotein strategy when translating its mRNA? Why aren’t the individual viral genes translated separately into the individual proteins?

Answer 38

because in eukaryotes (as a host cell) they are monocistronic, so they can only read 1 gene and stop, but the viral RNA is polycistronic, meaning that the eukaryotes ribosomes can't read all of them, so the poliovirus creates a large polyprotein that cleaves itself into smaller units.

Question 39

What is VPg and why is it important? If the gene encoding VPg was deleted, what effect would this have on the virus replication cycle?

Answer 39

VPg is a protein encoded by virus genome It is important because it acts as a primer for replication, by having 2 uracils and a 3'OH group add to it. Without it, no replication happens because the viral RNA polymerase cannot add new nucleotides

Question 40

In a laboratory experiment, human cells were infected with poliovirus and a mutant version of poliovirus is isolated. In this mutant, the entire 5’ UTR region of the genome is deleted. When this mutant virus is used to infect new host cells, what process(es) will be affected by this deletion? What will the effect be on the virus replication cycle?

Answer 40

there will no structural genes, so there will be no capsid proteins like VP0, VP1, VP3, so the virus cannot assemble. AND 5' UTR forms IRES that attracts ribosomes, without it, there will be no translation