CSB351Y1 Lecture Notes - Lecture 38: Polyomaviridae, Dna Gyrase, Sv40
26 May 2018
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Lecture 38: Virus with small DNA genomes
• Papovaviridae
- Polyomaviridae (small circular dsDNA):
◼ Simian virus 40 (SV40) – from mice with leukemia extracts
• Polyomavirus – naked (unenveloped) icosahedral capsid, 45nm
- Have histones forming nucleosomes (DNA arrangement)
- Can remove histones with phenol → supercoiled DNA → induce cleavage for loosening and ligase or
topoisomerase → DNA gyrase (recoil into supercoiled)
Genome
• 5kb and encodes protein – agnoprotein that enhances assembly of virus particles and cell to cell spread
• Divergent transcription – ½ ORF pointing at one direction ½ ORF at opposite direction (early vs late gene)
- Early – ST-Ag (small T antigen), LT-Ag (large T), MT-Ag and TT-Ag (tiny T) with SELP promoter
- Late – VP1,2,3,4 (capsid proteins) with AGNO promoter
• ATPase/helicase – transcription activation; origin binding domain (OBD) – DNA replication, LXCXE – cell cycle
control; J – transformation/making cancer
- This is inside the T antigens
• Large T Ag – activates/suppresses transcription of several cellular genes
• Small T Ag – induces cell division
• Middle T Ag – induces cell division (in different ways)
Events in SV40 virus replication
1. Primary (early) transcription by host enzymes
- Virus enters cell, endocytosed and releases DNA into nucleus
2. Translation of early (regulatory) proteins – T antigen genes transported out of cell and early translation
3. Viral genomic DNA replication
- Once sufficient, T antigen goes back into nucleus to aid with DNA replication (large T antigen)
4. Late transcription
- Cellular polymerase bind to late promoter and synthesize capsid proteins
5. Synthesis of late (structural) protein
6. Assembly into virions
Cellular RNA polymerase II transcribes T antigens
• Keeps cells in S (synthesis) phase so that cells can express DNA polymerase, proteins involved in DNA replication
• Can suppress cellular transcription
• LT-Ag forms hexamer, binds to viral ori and acts as helicase to unwind two DNA strands
- Assembles cellular DNA synthesis machinery initiating viral DNA replication
SV40 DNA replication
1. T-Ag binds to ori as double hexamer
2. T-Ag catalyzes melting of DNA duplex
3. Recruits cellular DNA polymerase onto origin
4. T-Ag acts as replicative helicase that unwinds DNA in front of the replication fork
• Small virus genome relies heavily on host cell machinery
find more resources at oneclass.com
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Document Summary
Lecture 38: virus with small dna genomes: papovaviridae. Simian virus 40 (sv40) from mice with leukemia extracts: polyomavirus naked (unenveloped) icosahedral capsid, 45nm. Can remove histones with phenol supercoiled dna induce cleavage for loosening and ligase or topoisomerase dna gyrase (recoil into supercoiled) Genome: 5kb and encodes protein agnoprotein that enhances assembly of virus particles and cell to cell spread, divergent transcription orf pointing at one direction orf at opposite direction (early vs late gene) Early st-ag (small t antigen), lt-ag (large t), mt-ag and tt-ag (tiny t) with selp promoter. Late vp1,2,3,4 (capsid proteins) with agno promoter: atpase/helicase transcription activation; origin binding domain (obd) dna replication, lxcxe cell cycle control; j transformation/making cancer. Large t ag activates/suppresses transcription of several cellular genes. This is inside the t antigens: small t ag induces cell division, middle t ag induces cell division (in different ways) Events in sv40 virus replication: primary (early) transcription by host enzymes.
