RNA VIRUS REPLICATION/ TRNSLATION STRATEGIES Sadia Anjum Virology lecture 5 1.
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Transcript of RNA VIRUS REPLICATION/ TRNSLATION STRATEGIES Sadia Anjum Virology lecture 5 1.
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RNA VIRUS REPLICATION/ TRNSLATION STRATEGIES
Sadia Anjum
Virology lecture 5
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RNA VIRUS STRATEGIES
RNA -> RNARNA-dependent RNA polymerase
RNA -> DNARNA-dependent DNA polymerase- reverse transcriptase
Host cell DNA -> RNADNA-dependent RNA polymerase
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All animal RNA viruses code for a polymerase
Need to make mRNAPLUS (POSITIVE) SENSE RNA GENOMES
AAA
(+ve) sense mRNA
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NEED TO MAKE MRNA
MINUS (NEGATIVE) SENSE RNA GENOMES
AAA(+ve) sense mRNA
(-ve) sense genomic RNA
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NEED TO MAKE MRNAMINUS (NEGATIVE) SENSE RNA GENOMES
RNA polymerase must be packaged in virion.
AAA(+ve) sense mRNA
(-ve) sense genomic RNAIf used, RNA modifying enzymes arepackaged in virion.
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DOUBLE-STRANDED RNA GENOMES
AAA(+ve) sense mRNA
Double-stranded genomic RNAIf used, RNA modifying enzymes arepackaged in virion.
RNA polymerase must be packaged in virion.
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RETROVIRUSES
DSDNA
+VE RNA
DSDNA
Reverse transcriptase must be packaged in virion.
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RNA VIRUSES THAT DO NOT HAVE A DNA PHASE
TranscriptionNon-infectiousYesDouble-stranded RNA
TranscriptionNon-infectiousYesNegative-stranded RNA
TranslationInfectiousNoPlus-stranded RNA
Initial event in cell
Infectivity of RNA
RNA-dependent RNA polymerase
(=transcriptase) IN VIRION
Genome
TranscriptionNon-infectiousYesDouble-stranded RNA
TranscriptionNon-infectiousYesNegative-stranded RNA
TranslationInfectiousNoPlus-stranded RNA
Initial event in cell
Infectivity of RNA
RNA-dependent RNA polymerase
(=transcriptase) IN VIRION
Genome
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RETROVIRUSESGenome RNA-
dependent DNA polymerase (=reverse transcriptase) in virion
Infectivity of RNA
Initial event in cell
Plus-stranded RNA
Yes Non-infectious Reverse transcription
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THE MONOCISTRONIC MRNA PROBLEM
Make one monocistronic mRNA per protein Make a primary transcript and use alternative splicing Make a large protein and then cut it into smaller
proteins Include special features in the mRNA which enable
ribosomes to bind internally
AAAAAAAA
RIBOSOMES
mRNA
PROTEIN
AAAAAAAA
Eukaryotes
Prokaryotes/viruses
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POLYCISTRONIC VS MONOCISTRONIC
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RNA GENOME SIZE
TAUTOMERIZATION PROOF READING GENOME SIZE
RNA VIRUSES HAVE SMALL GENOMES~10,000 NUCLEOTIDES(herpesvirus DNA ~200,000)
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GENOME SIZE POLYMERASE ATTACHMENT PROTEIN CAPSID PROTEIN RELATIVELY FEW OTHER PROTEINS VIRION SIZE MAY BE QUITE LARGE BUT
LIMITED NUMBER OF PROTEINS
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POSITIVE-STRANDED RNA VIRUSES
EXAMPLES
PICORNAVIRUSES
TOGAVIRUSES
FLAVIVIRUSES
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PICORNAVIRUS FAMILY (PICORNAVIRIDAE)
SMALL ICOSAHEDRAL POSITIVE SENSE RNA NON-ENVELOPED
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INTERNAL RIBOSOME ENTRY SITE (IRES)
GENOMIC (+ SENSE) RNAAAAAAVPg
IRES
start codonfor translation
stop codonfor translation
POLYPROTEIN
.
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VPG VPg (viral protein genome-linked) is a protein that is
covalently attached to the 5′ end of positive strand viral RNA and acts as a primer during RNA synthesis in a variety of virus families including Picornaviridae and Caliciviridae.
The primer activity of VPg occurs when the protein becomes uridylylated, providing a free hydroxyl that can be extended by the virally encoded RNA-dependent RNA polymerase.
Aside from functioning as a primer, VPg also has a role in translation initiation by acting like a 5' mRNA cap
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INHIBITION OF HOST CELL TRANSLATION
HOST mRNAAAAAACAP
Uninfected cell
Factors+40S ribosomal subunit
HOST mRNAAAAAACAP
Altered factors+40S ribosomal subunit
Infected cell
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VIRAL TRANSLATION
GENOMIC (+ SENSE) RNAAAAAA
IRES
VPg
Factors+40S ribosomal subunit
Altered factors+40S ribosomal subunit
GENOMIC (+ SENSE) RNAAAAAA
IRES
VPg
Uninfected cell
Infected cell
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GENOMIC (+ SENSE) RNAAAAAA
translation
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RNA REPLICATION
GENOMIC (+ SENSE) RNA 3’VPg
(- SENSE) RNA 3’ VPg
GENOMIC (+ SENSE) RNA 3’VPg
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RNA REPLICATION viral RNA polymerase (replicase) host factors also involved as accessory
proteins new plus strands
packaged templates for more replication templates for more translation
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ASSEMBLY
VP0, VP1, VP3
VP2, VP4, VP1, VP3VPg
VPg-RNA enters, VP0 is cleaved
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NON-SEGMENTED NEGATIVE STRAND VIRUSES
Examples:
Rhabdovirus family (Rhabdoviridae)
Paramyxovirus family (Paramyxoviridae)
Filovirus family (Filoviridae)
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RHABDOVIRUSES
helical nucleocapsid (RNA plus N protein)
G glycoprotein SPIKES
lipid bilayer membrane
polymerase complex
M protein
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MINUS (NEGATIVE) SENSE RNA GENOME
RNA polymerase must be packaged in virion.
AAA(+ve) sense mRNA
(-ve) sense genomic RNA
RNA modifying enzymes are packaged in virion.
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RHABDOVIRUSES
EXAMPLESvesicular stomatitis
virus (VSV)
rabies virus
Dr FS Murphyhttp://www.vetnet.ucdavis.edu/fam_graphics/download.html
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ADSORPTION AND PENETRATION
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CYTOPLASMIC REPLICATION
GENOMIC RNA REMAINS IN NUCLEOCAPSID FORM
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RNA SYNTHESIS
3’ 5’genome (-ve)
3’ 5’ (-ve)
full-length copy
5’ 3’ (+ve)
replication (protein synthesis is a pre-requisite)
transcription
mRNAs (+ve sense)
AAA
AAA
AAA
AAA
AAA
= cap
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NEW MINUS STRAND
PACKAGED TEMPLATE FOR
MORE REPLICATION TEMPLATE FOR
MORE TRANSCRIPTION
3’ 5’genome (-ve)
3’ 5’ (-ve)
5’ 3’ (+ve)
AA
AA
AA
AA
AA
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SOME POINTS TO NOTE ABOUT RHABDOVIRUSES
ENTIRE CYCLE OCCURS IN CYTOPLASM
RNA POLYMERASE AND RNA MODIFICATION
ENZYMES ARE:
VIRALLY CODED
PRESENT IN THE VIRION
THERE IS NO EARLY/LATE DIVISION
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MEMBERS INCLUDE
PARAINFLUENZA VIRUS
MUMPS VIRUS
MEASLES VIRUS
RESPIRATORY SYNCYTIAL VIRUS
PARAMYXOVIRUSES
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PARAMYXOVIRUSES
M protein
helical nucleocapsid (RNA plusNP protein)
HN/H/G glycoprotein SPIKES
polymerase complex
lipid bilayer membrane
F glycoprotein SPIKES
pleomorphic
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PARAMYXOVIRIDAE
PARAMYXOVIRUS SUBFAMILY
PARAMYXOVIRUS
RUBULAVIRUS
MORBILLIVIRUS
PNEUMOVIRUS SUBFAMILY
PNEUMOVIRUS GENUS
METAPNEUMOVIRUS GENUS
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GENUS GLYCOPROTEINS TYPICAL MEMBERS
Paramyxovirus HN, F HPIV1, HPIV3Rubulavirus HN, F HPIV2, HPIV4, mumps virusMorbillivirus H, F measles virus
Pneumovirus G, F respiratory syncytial virusMetapneumovirus G, F metapneumoviruses
PARAMYXOVIRUS FAMILY SURFACE GLYCOPROTEINS
PARAMYXOVIRUS SUBFAMILY
PNEUMOVIRUS SUBFAMILY
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TRANSCRIPTION, TRANSLATION, REPLICATION
3’ 5’genome (-ve)
full-length copy
5’ 3’ (+ve)
3’ 5’ (-ve)
replication (protein synthesis is a pre-requisite)
transcription
mRNAs (+ve sense)
AAA
AAA
AAA
AAA
AAA
= cap
AAA
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SOME DIFFERENCES BETWEEN RHABDOVIRUSES AND PARAMYXOVIRUSES
two:
one attachment
one fusion
one (has both attachment and fusion activities)
glycoproteins
neutral
physiological
acidicfusion pH
round
pleomorphic
bullet
bacilliform
shape
PARAMYXOVIRUSESRHABDOVIRUSES
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ORTHOMYXOVIRUSES
M1 protein
helical nucleocapsid (RNA and NP protein)
HA - hemagglutinin
polymerasecomplex
lipid bilayer membrane
NA - neuraminidase
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ADSORPTION AND PENETRATION
NUCLEUS
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TRANSCRIPTION (MRNA SYNTHESIS)
AAAA
viral endonuclease
A
host cell mRNA
influenza mRNA
=cap
influenza virus nucleocapsidAU
AU
viral polymerase
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RNA SYNTHESIS
Endonuclease is virally coded and packed in the
virion
RNA polymerase is virally coded and packed in
the virion
Poly(A) polymerase is virally coded and in the
virion
Why doesn’t the virus use host cell enzymes?
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RNA SPLICING
Several segments give rise to transcripts which can have one of two fates
AAAAAA
AAAAAA
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RNA REPLICATION nuclear
authentic copies
replicase and
transcriptase probably
same enzyme
coated with protein as
made
no clear early/late
3’ 5’genomic (-ve)
3’ 5’ genomic (-ve)
5’ 3’ (+ve)
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NUCLEUS
acid pH inside endosome
- HA conformational change, now active in membrane fusion
Infection of next cell
HA cleavage by host cell enzymes
Flu Virus
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PARAMYXOVIRIDAE ORTHOMYXOVIRIDAE
genome nonsegmented segmented
RNA synthesis cytoplasmic nuclear
need for RNA primer
no yes
hemagglutinin, neuraminidase
if both, part of same protein (HN)
Influenza A and B viruses have both, but on different proteins (HA and NA)
syncytial formation
yes (F functions at physiol. pH)
no (HA functions at acid pH)
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REOVIRUS FAMILYdouble stranded RNA genome
outer capsid
inner capsid
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DOUBLE-STRANDED RNA GENOMES
AAA(+ve) sense mRNA
Double-stranded genomic RNARNA modifying enzymes arepackaged in virion.
RNA polymerase must bepackaged in virion.
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REOVIRUS FAMILYdouble stranded RNA genome
outer capsid
inner capsid
RNA polymerase plus 5’ endmodification enzymes
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REOVIRUS FAMILY INCLUDES
REOVIRUSES ROTAVIRUSES
IMPORTANT HUMAN PATHOGENS ORBIVIRUSES
INCLUDE COLORADO TICK FEVER VIRUS
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Annual Review of Microbiology 51:227 RF Ramig