RNA VIRUSESRNA VIRUSES
ALL SORTS OF STRATEGIES
RNA VirusesRNA Viruses• All synthesize through a
double stranded intermediate - RI - replication intermediate
• RNA dependent RNA polymerase of viral origin but may need host factors
• Termini contain recognition signals for replicase
Positive strand virusesPositive strand viruses
• Begin with translation to produce replicase
• Makes more positive than negative strand
– Limiting factor or rapid packaging so can’t act as template
• Poliovirus uses VPg linked to nucleotides as “primer” - like Ad
Negative Strand VirusesNegative Strand Viruses
• Contain enzymes for transcription in virion• Make mRNA prior to antigenome
– Message gets capped; genome does not• Plus strand is template for minus strand genome• Makes more minus than plus strand
dsRNA viruses - conservative dsRNA viruses - conservative replicationreplication
• Uncoating activates enzymes that produce mRNA
• + RNA also gets packaged
• Then complementary - RNA is produced
• No dsRNA free in cell
• Protects against IF induction
Transcription challengesTranscription challenges• Less temporal control than
in DNA viruses• Monogenic problem
– Segmented genomes usually have individual genes
– Polyprotein cleavage• What would expect to see
on gel in early stages of infection? As infection progresses?
• What if you performed a pulse-chase experiment?
TranslationTranslation challengeschallenges:: Recognition by ribosomes Recognition by ribosomes and competition from hostand competition from host
• Synthesize own cap (Reo in cytoplasm)
• Steal from host (Influenza in nucleus)
• Use host enzymes
• IRES
Transcription strategies: TogavirusesTranscription strategies: Togaviruses
• NS at 5’ end - S at 3’• In vitro only synthesize
NS proteins; stop signal leads to polyprotein
• In vivo get shorter mRNA only after minus strand synthesis that codes for S polyprotein
• Internal transcription site on minus strand
• Minus is template for mRNA and for genome
• S message is more abundant than NS as genome gets packaged
Coronaviruses: frame-shifts and subgenomic Coronaviruses: frame-shifts and subgenomic RNAsRNAs
• Genome translated into replicase
• Antigenome produced• Subgenomic mRNAs represent
a nested set of RNAs - all share short 5’ sequence and a 7 base sequence but have unique AUG site and share 3’ end of genome
• May be produced by jumping polymerase - 7 base sequence in various parts of genome– Get recombinant viruses
with mixed infections– DI particles are common
Influenza virus - segmented negative strandInfluenza virus - segmented negative strandantigenic drift (mutations) vs shift (reassortment)antigenic drift (mutations) vs shift (reassortment)
Influenza - negative strand virusInfluenza - negative strand virus
• Replication in nucleus using viral enzymes but need host RNA-P to function
• Virion enzyme cleaves cap from host mRNA and uses it to extend; adds poly A tail
• One gene per segment except for two segments producing spliced mRNAs in two different reading frames yielding two proteins
Ambisense genomesAmbisense genomes
• Bunyaviridae such as Hantavirus
• Genome is used to make short positive mRNA
• Genome is replicated and antigenome (plus strand) is used to make second mRNA
• Antigenome does not act as message
Nonsegmented negative strand viruses: Nonsegmented negative strand viruses: Mononegavirales (rhabdo, filo, paramyxo)Mononegavirales (rhabdo, filo, paramyxo)
• Hypothesis: Start-stop
• Template 3’ end start point for virion L (RNA_P) and goes to termination signal and mRNA release - then cap and poly A added
• Some polymerase reinitiates at next initiation signal and goes to termination; process repeats
• Each subsequent RNA may be produced at a lower frequency (20-30% less)
• Replication requires N capsid and NS proteins to read through to complete copy
Retroviruses: diploid ssRNA with Retroviruses: diploid ssRNA with repeats at endsrepeats at ends
• RT needs a primer - uses tRNA at primer binding site
• Synthesized to end and jumps to 3’ end of strand
• Uses PPT as template for second strand
• Makes another jump• Results in dsDNA with Long
Terminal Repeats– Needed for integration– Contains promotor and
regulatory regions– Poly A site
Transcription occurs after integrationTranscription occurs after integration
• Uses host RNA-P
• May require host factors to enhance (cell tropism)
• Polyprotein and splicing strategies
HIV is a more complex retrovirusHIV is a more complex retrovirus
• Transactivator protein (TAT) needed for high level of transcription
• TAT binds to TAR RNA and causes readthrough beyond 5’ region
REV binds to REV Response Element REV binds to REV Response Element (RRE) in message(RRE) in message
• Early messages are highly spliced and produce mainly TAT, REV and Nef
• When REV increases and binds to message, there is less splicing
• Leads to synthesis of gag, pol, env
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