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Lecture 8 - (+)-Sense RNA Viruses Flashcards

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1
Q

Baltimore class of most (+)ssRNA viruses

A

Class IV

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2
Q

(+)RNA viral families

A

1) Caliciviridae 2) Picornaviridae 3) Togaviridae 4) Flaviviridae 5) Retroviridae (Baltimore VI, not IV) 6) Coronaviridae

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3
Q

Features of viral (+)ssRNA 1) 2) 3) 4) 5) 6)

A

1) Possible 5’ m7G cap 2) Possible poly-A tail 3) Sequence editing 4) Have no introns 5) Translationally active 6) Resist degradation

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4
Q

(+)ssRNA viruses with 5’ m7G cap and poly-A tail

A

Coronaviridae Togaviridae

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5
Q

(+)ssRNA viruses with a 5’ m7G cap

A

Coronaviridae Togaviridae Flaviviridae

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6
Q

(+)ssRNA viruses with a poly-A tail

A

Coronaviridae Togaviridae Picronaviridae

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7
Q

(+)ssRNA virus without a 5’ m7G cap

A

Picornaviridae

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8
Q

(+)ssRNA virus without a poly-A tail

A

Flaviviridae

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9
Q

Only (+)ssRNA virus that carries a capsid polymerase

A

Retroviridae (carry reverse transcriptase)

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10
Q

Why is the cytoplasm a hostile place for a virus? 1) 2)

A

1) dsRNA in the cytoplasm is detected, induces IFN secretion 2) Little machinery for genome replication

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11
Q

Gene in coronaviridae, flaviviridae, togaviridae for methyl cap

A

Methyl transferase enzyme

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12
Q

Essential protein for (+)ssRNA replication

A

RNA-dependent RNA polymerase (virus encoded)

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13
Q

ssRNA replication 1) 2) 3) 4)

A

1) Requires RNA-dependent RNA polymerase 2) Can copy both (+) and (-) ssRNA 3) Complex is partly double-stranded (called a replicative intermediate) 4) Normally, few (-) strands are made, and many (+) strands are replicated from the (-) strands

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14
Q

Viruses that make a single RNA

A

Picornaviridae Flaviviridae

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15
Q

Viruses that make subgenomic RNA

A

Togaviridae Caliciviridae

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16
Q

Examples of picronaviridae

A

Poliovirus Hepatitis A virus Rhinovirus

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17
Q

Examples of flaviviridae

A

West Nile/Dengue virus Hepatitis C virus

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18
Q

Examples of togaviridae

A

Rubellavirus Ross River virus Sindbis

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19
Q

Examples of calicivirus

A

Norovirus

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20
Q

Single mRNA method

A

Viral (+) genome replicates to a (-) genome, which is replicated to a (+) RNA. This (+) RNA is used for both translation and as a genome

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21
Q

Subgenomic RNA method

A

Viral genome is replicated into a (-) full genome. (-) genome is replicated to either a full genome or a subgenomic mRNA, which is used for protein translation

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22
Q

Poliovirus characteristics

A

Small, (+) ssRNA virus with a naked icosahedral capsid

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23
Q

What does the poliovirus genome encode?

A

A single polyprotein

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24
Q

Structure of poliovirus polyprotein

A
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25
Sections of polio polyprotein 1) 2) 3)
1) 5' end (P1) is structural proteins 2) P2 encodes proteins that affect the host (stop secretory system, proteases prevent host translation) 3) P3 encodes replicative genes
26
What cleaves polio polyprotein?
Viral proteases (2Apro, 3Cpro)
27
What is attached to the 5' end of polio RNA?
VPg protein
28
Poliovirus replication 1) 2) 3) 4)
1) VPg protein removed from genome when genome is inserted into cell 2) Polyprotein translated, P2, P3 proteases, enzymes, VPg move to a cytoplasmic membrane-bound vesicle. 3) VPg acts as a primer for RNA-dependent RNA polymerase (3Dpol) replication of (+) and (-) ssRNA 4) Particles assemble in membrane-bound vesicle, are released when cell is lysed
29
Poliovirus RNA-dependent RNA polymerase
3Dpol
30
Major issue for (+)ssRNA viruses
Translation occurs 5' -\> 3' Replication occurs 3' -\> 5' along (+) strand (assembles (-)RNA 5' -\> 3') Polymerase and ribosome will collide
31
Poliovirus RNA replication 1) 2) 3) 4) 5) 6) 7)
1) At the 5' end of RNA genome is a clover leaf structure. Host PCBP2 and viral 3CD bind this. 2) Host PABP binds 3' poly-A tail. PCBP2 binds to PABP, forming a loop of RNA. 3) PCBP2 and PABP interactions lead to 3CD cleavage into 3C and 3D-RNA-dependent RNA polymerase. 4) 3D begins replication of genome, utilising VPg-UUU sequence as a primer. 5) Extension of (-) RNA 6) Host chaperone hnRNPC binds ends of (-)RNA. hnRNPC binding to (-)RNA and PCBP2/PABP binding to (+) lead to partial unwinding of dsRNA 7) (+)RNA synthesis occurs from unwound part.
32
How does poliovirus prevent collision of replication and translation complexes on genomic RNA? 1) 2) 3)
1) A loop of RNA is formed by host PCBP2 binding to 5' clover leaf structure and host PABP binding to 3' poly-A tail 2) This loop prevents the access of ribosomes to (+)RNA. 3) Protein synthesis only occurs when the number of genomes outnumbers the number of PCBP2/PABP.
33
How does poliovirus stop cellular cap-dependent translation?
Polio 2A protease cleaves cellular elF4-G, which is part of the complex that initiates cap-dependent translation
34
Effect of polio 2A protease cleaving cellular elF4-G
Prevents cellular ability to initiate translation of mRNAs with a cap. This allows only viral proteins to be translated in a cell
35
How long does it take poliovirus to shut down host-cell translation?
About 2 hours
36
How does polioviral mRNA initiate translation?
5' portion folds into an internal ribosome entry site (IRES). Binds directly to cellular translation factors
37
Characteristic structures in a late-stage poliovirus-infected cell
Double-membrane bound vesicles. Contain dividing virus
38
How does poliovirus evade IFN production in response to dsRNA detection?
Viral genome replication occurs in vesicles within the cytoplasm.
39
Why does poliovirus need to replicate genomes within membrane-bound vesicles?
An intermediate stage of viral genome replication is dsRNA. This is detected by a cell and is a powerful stimulus for IFN release
40
Differences in how hepatitis C virus and poliovirus initiate translation
HCV - Has an internal ribosome entry site (IRES) on the 5' end Poliovirus - VPg-like protein and an internal ribosome entry site
41
Does HCV directly kill hepatocytes?
No. Unlike poliovirus, it doesn't interfere with host cell translation
42
Togaviridae encoded proteins
Two polyproteins: P123 and P1234
43
Virus that encodes P123 and P1234
Togaviridae
44
P123 and P1234 differential expression
Togaviruses have a slippery stop codon that can be read through in their genomes. If translation terminates with the stop codon, P123 results. If the stop codon is read through, P1234 results.
45
Togaviridae genome replication 1) 2) 3) 4) 5)
1) P123 and P1234 are translated 2) P1234 polyprotein is cleaved by viral protease nsP2. This forms enzymes needed for viral genome replication 3) A (-)ssRNA copy of the genome is made. 4) Two (+)ssRNA copies of the (-)ssRNA genome are made: a genome-length RNA and a subgenomic RNA 5) Subgenomic RNA encodes structural proteins
46
How do togaviruses regulate when structural proteins are expressed?
1) The subgenomic promotor region only transcribes from (-)ssRNA, which only appears after P123 has been cleaved into enzymes needed for viral replication. Therefore it is initially 'hidden' from the host ribosome
47
Viral family that produces subgenomic RNAs from 3' end of genome
Togaviridae
48
How do Togaviridae regulate when structural genes are translated? 1) 2) 3)
1) Capsid genes are at the 3' end in Togaviridae 2) Togaviridae can produce subgenomic RNAs from the 3' end of genome. 3) This means that only when replication has been established can capsid proteins be made (as a lot of (-)RNA is needed
49
How do togaviridae regulate whether full-length RNA or subgenomic RNA are expressed? 1) 2) 3) 4)
1) Progressive proteolytic cleavage of RNA-replicase changes enzyme preference for which RNA to transcribe 2) First cleavage activates (-)RNA synthesis from (+)RNA template 3) Second cleavage activates full genome (+)RNA synthesis from (-)RNA template 4) Full proteolytic cleavage activates subgenomic (+)mRNA for structural proteins from full-length (-)RNA template
50
SARS coronavirus features 1) 2) 3) 4)
1) Irregularly-shaped enveloped virion 2) Very large (27-31kb) genome 3) Helical nucleocapsid 4) No virion polymerase
51
Only known helical nucleocapsid with (+)ssRNA
SARS coronavirus
52
Structure of a coronavirus
E2 - Spike glycoprotein HA - Haemagglutinin (also called E3) E1 - Membrane glycoprotein (also called M) N - Nucleoprotein Es - Envelope glycoprotein
53
Spike glycoprotein of SARS
E2
54
Membrane glycoprotein of SARS
E1
55
Envelope glycoprotein of SARS
Es
56
Virus with a 'nested set' of mRNAs
SARS
57
SARS replication 1) 2) 3) 4) 5) 6) 7)
1) Enters cell by fusion 2) RNA-dependent RNA polymerase translated fmo genomic RNA 3) (-)ssRNA replicative intermediate is transcribed in the cytoplasm 4) 5' leader end of the (+)RNA is transcribed with a 5' methyl cap, added by viral enzyme 5) RNA polymerase skips to downstream sites to make a 'nested set' of non-identical mRNAs 6) Viral particles bud into the Golgi 7) Infectious particles are released from the plasma membrane
58
Formation of 'nested sets' of SARS mRNA 1) 2) 3) 4)
1) Genome-length mRNA has a methyl cap added by viral polymerase 2) Common leader RNA sequence is joined to one of many repeated intergenic sequences (UCUAAAC). 3) This leads to the RNA-dependent RNA polymerase to skip the sequences between the common leader RNA sequence and the intergenic sequence 4) This produces a set of 8 discontinuous subgenomic mRNAs
59
Sequence of repeated intergenic sequence in SARS genome
UCUAAAC
60
Different locations of structural genes on Picornaviruses and Togaviruses
Structural genes on 5' end of Picornaviridae genome, on 3' end of Togaviridae genome

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