DNA-dependent RNA polymerase 2 All animal RNA viruses code for a polymerase Need to make mRNA PLUS (POSITIVE) SENSE RNA GENOMES proteins AAA (+ve) sense mRNA 3 NEED TO MAKE MRNA
MINUS (NEGATIVE) SENSE RNA GENOMES proteins (+ve) sense mRNA AAA (-ve) sense genomic RNA 4 NEED TO MAKE MRNA MINUS (NEGATIVE) SENSE RNA GENOMES RNA polymerase must be packaged in virion. proteins (+ve) sense mRNA If used, RNA modifying
enzymes are packaged in virion. AAA (-ve) sense genomic RNA 5 DOUBLE-STRANDED RNA GENOMES RNA polymerase must be packaged in virion. proteins (+ve) sense mRNA If used, RNA modifying enzymes are packaged in virion.
AAA Double-stranded genomic RNA 6 RETROVIRUSES Reverse transcriptase must be packaged in virion. DS DNA DS DNA +VE RNA 7
RNA VIRUSES THAT DO NOT HAVE A DNA PHASE Genome RNA-dependent Infectivity of RNA RNA polymerase (=transcriptase) IN VIRION Initial event in cell Plus-stranded RNA No
RETROVIRUSES Genome Plus-stranded RNA RNAdependent DNA polymerase (=reverse transcriptase) in virion Infectivity of RNA Initial event in cell
Yes Non-infectious Reverse transcription 9 THE MONOCISTRONIC MRNA PROBLEM mRNA AAAAAAAA Eukaryotes
RIBOSOMES PROTEIN AAAAAAAA Prokaryotes/viruses 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 10 11
POLYCISTRONIC VS MONOCISTRONIC 12 RNA GENOME SIZE TAUTOMERIZATION PROOF READING GENOME SIZE RNA VIRUSES HAVE SMALL GENOMES ~10,000 NUCLEOTIDES (herpesvirus DNA ~200,000) 13
GENOME SIZE POLYMERASE ATTACHMENT PROTEIN CAPSID PROTEIN RELATIVELY FEW OTHER PROTEINS VIRION SIZE MAY BE QUITE LARGE BUT LIMITED NUMBER OF PROTEINS 14 POSITIVE-STRANDED RNA VIRUSES EXAMPLES
PICORNAVIRUSES TOGAVIRUSES FLAVIVIRUSES 15 PICORNAVIRUS FAMILY (PICORNAVIRIDAE) SMALL ICOSAHEDRAL
POSITIVE SENSE RNA NON-ENVELOPED 16 INTERNAL RIBOSOME ENTRY SITE (IRES) IRES GENOMIC (+ SENSE) RNA VPg POLYPROTEIN start codon for translation
. AAAAA stop codon for translation 17 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 RNAdependent RNA polymerase. Aside from functioning as a primer, VPg also has a role in translation initiation by acting like a 5' mRNA cap 18
INHIBITION OF HOST CELL TRANSLATION Uninfected cell CAP HOST mRNA AAAAA Factors+40S ribosomal subunit Infected cell CAP HOST mRNA AAAAA
3 23 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 24
ASSEMBLY VP0, VP1, VP3 VPg-RNA enters, VP0 is cleaved VPg VP2, VP4, VP1, VP3 25 NON-SEGMENTED NEGATIVE STRAND VIRUSES Examples: Rhabdovirus family (Rhabdoviridae) Paramyxovirus family (Paramyxoviridae) Filovirus family (Filoviridae)
26 RHABDOVIRUSES G glycoprotein SPIKES M protein lipid bilayer membrane helical nucleocapsid (RNA plus N protein) polymerase complex 27 MINUS (NEGATIVE) SENSE RNA GENOME RNA polymerase must be packaged in virion. RNA modifying enzymes are packaged in virion.
proteins (+ve) sense mRNA AAA (-ve) sense genomic RNA 28 RHABDOVIRUSES EXAMPLES vesicular stomatitis virus (VSV) rabies
virus Dr FS Murphy http://www.vetnet.ucdavis.edu/fam_graphics/download.html 29 ADSORPTION AND PENETRATION H+ 30 CYTOPLASMIC REPLICATION
GENOMIC RNA REMAINS IN NUCLEOCAPSID FORM 31 RNA SYNTHESIS genome (-ve) 3 replication (protein synthesis is a pre-requisite) transcription mRNAs (+ve sense) AAA AAA
5 full-length copy AAA 5 (+ve) 3 AAA AAA 3 = cap
(-ve) 5 32 NEW MINUS STRAND PACKAGED FOR MORE REPLICATION TEMPLATE FOR MORE TRANSCRIPTION 3 genome (-ve)
5 TEMPLATE AA AA 5 (+ve) 3 AA AA
AA 3 (-ve) 5 33 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 34 PARAMYXOVIRUSES
MEMBERS INCLUDE PARAINFLUENZA MUMPS VIRUS VIRUS MEASLES VIRUS RESPIRATORY SYNCYTIAL VIRUS 35
PARAMYXOVIRUSES pleomorphic HN/H/G glycoprotein SPIKES F glycoprotein SPIKES helical nucleocapsid (RNA plus NP protein) lipid bilayer membrane polymerase complex M protein 36 PARAMYXOVIRIDAE
PARAMYXOVIRUS SUBFAMILY PARAMYXOVIRUS RUBULAVIRUS MORBILLIVIRUS PNEUMOVIRUS SUBFAMILY PNEUMOVIRUS GENUS METAPNEUMOVIRUS GENUS 37
PARAMYXOVIRUS FAMILY SURFACE GLYCOPROTEINS GENUS GLYCOPROTEINS PARAMYXOVIRUS SUBFAMILY Paramyxovirus Rubulavirus Morbillivirus PNEUMOVIRUS SUBFAMILY Pneumovirus Metapneumovirus HN, F HN, F
H, F G, F G, F TYPICAL MEMBERS HPIV1, HPIV3 HPIV2, HPIV4, mumps virus measles virus respiratory syncytial virus metapneumoviruses 38 TRANSCRIPTION, TRANSLATION, REPLICATION genome (-ve) 3
replication (protein synthesis is a pre-requisite) transcription mRNAs (+ve sense) AAA AAA AAA 5 5 full-length copy (+ve)
3 AAA AAA AAA = cap 3 (-ve) 39 5 SOME DIFFERENCES BETWEEN RHABDOVIRUSES AND PARAMYXOVIRUSES
RHABDOVIRUSES PARAMYXOVIRUSES shape bullet bacilliform round pleomorphic glycoproteins one (has both attachment and fusion activities) two: one attachment one fusion fusion pH
neutral physiological acidic 40 ORTHOMYXOVIRUSES HA - hemagglutinin NA - neuraminidase helical nucleocapsid (RNA and NP protein) lipid bilayer membrane polymerase complex 41
M1 protein NUCLEUS ADSORPTION AND PENETRATION H+ 42 TRANSCRIPTION (MRNA SYNTHESIS) =cap host cell mRNA A AAA
viral endonuclease A A U influenza virus nucleocapsid viral polymerase A U influenza mRNA 43 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 doesnt the virus use host cell enzymes?
44 RNA SPLICING Several segments give rise to transcripts which can have one of two fates AAAAAA AAAAAA 45 RNA REPLICATION nuclear
authentic copies replicase and 3 genomic (-ve) 5 transcriptase probably same enzyme coated with protein as
5 3 (+ve) made no clear early/late genomic (-ve) 3 5 46
NUCLEUS Flu Virus Infection of next cell HA cleavage by host cell enzymes H+ - HA conformational change, now active in membrane fusion 47
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