Virology II Lecture Notes (2023)

Lecture 37 Virology II Viral Replication Learning. Problems in Viral Replication viruses need RNA Replicase (RNA dependent RNA polymerase) viruses need Reverse Transcriptase (RNA dependent DNA poly  From last class: How might a plus sense ssRNA virus undergo re...

Lecture 37 Virology II Viral Replication Learning. Problems in Viral Replication viruses need RNA Replicase (RNA dependent RNA polymerase) viruses need Reverse Transcriptase (RNA dependent DNA poly  From last class: How might a plus sense ssRNA virus undergo replication? How might a plus sense ssRNA virus 1 undergo 3 2 replication? 4 5 Uses its genome as mRNA for protein synthesis Source: Viral Zone The Diversity of What is the origin of viruses? Viruses Subviral particles  Where do viruses come from? Outer space? They are cells that degenerated into parasites They are selfish genes They are the remnants of another type of organism that once lived on earth but is now extinct Outline 1. Lytic Replication Cycle 2. Lysogenic Replication Cycle 3. Retrovirus Replication Cycle 4. Problems in Viral Replication 1. Lytic Replication Cycle Counting phage 1. Lytic Replication Cycle Lytic Cycle 1. Lytic Replication Cycle ic cycle example: T4 Phage (linear dsDNA genome) 1. Lytic Replication Cycle he ‘end problem’ of replicating linear genomes 1. Lytic Replication Cycle Circular Permutation is used to solve the ‘end problem’ in T4 -Genomes are recombined end to end to form a concatemer -Concatemer inserted into capsid head until capsid head is full -Capsid holds ~103% of genome (duplication at the ends). Lysogenic Replication Cycle Temperate Phage and Lysogeny Temperate (lysogenic) phage: A virus that can undergo lysogeny in addition to the lytic cycle. Lysogeny: Phage integrates in host genome where it is replicated during host DNA replication and inherited by host progeny – Lytic pathway is inducible by inactivation of a phage repressor protein. Prophage: Viral DNA covalently integrated into host DNA.. Lysogenic Replication Cycle. Lysogenic Replication Cycle ysogenic Cycle Example: ambda Phage (linear dsDNA genome) cos site: cohesive ends, single ay w stranded overhang, allows for a th Lysogenic pathway p circularization t ic Ly Rolling circle replication: lytic pathway, genome synthesized and cut at cos site Lambda integrase: lysogenic pathway, phage endonuclease. Lysogenic Replication Cycle egulation of lysogeny in Lambda Phage 2 promoters: PRM (for cI) and PR (for cro) (cI) 3 operators: OR1, OR2, OR3 2 transcription factors: Lambda () repressor (cI) Lambda repressor (cI): suppresses lytic cro repressor pathway (cro) -represses cro promoter (PR) -activates cI promoter (PRM) -is cleaved by RecA (recall SOS response) Cro repressor: triggers lytic pathway http://www.amolf.nl/research/biochemical-networks/research-. Lysogenic Replication Cycle Theta replication: typical dual replication forks starting at ori Rolling circle replication: circular ssDNA template, one replication fork Lambda Activator II (CII): triggers lysogeny on infection, unstable protein, if conditions Blue = viral DNA favor protein stability then CII high – if cell is Green = viral mRNA  (cI) Which of the following might cause a lambda prophage to become ‘trapped’ in a bacterial genome? A. A deletion of the lambda repressor (cI) B. A non-sense mutation in the cro repressor C. A silent mutation in the lambda repressor D. A silent mutation in the lambda integrase gene E. Both C and D would trap a prophage within a bacterial genome. Retrovirus Replication Cycle Retrovirus Example: HIV (linear ssRNA (+) genome) Retrovirus: ssRNA (+) virus that uses a dsDNA intermediate which is inserted into the host genome Note: HIV has terminal repeats which it uses to solve the ‘end problem’ of. Retrovirus Replication Cycle Retrovirus Example: HIV (linear ssRNA (+) genome) HIV virion includes: ssRNA (+) genome, reverse transcriptase, integrase, and a tRNA-primer Reverse Transcriptase: a multifunctional enzyme that converts ssRNA into dsDNA, has the following activities: -RNA dependent DNA polymerase -DNA dependent DNA polymerase -ribonuclease (degrade RNA from RNA:DNA hybrid) Integrase: integrate dsDNA into host genome. Retrovirus Replication Cycle Retrovirus Example: HIV (linear ssRNA (+) genome) everse Transcriptase (RT): Integrase i) tRNA primes RT ii) RNA dependent DNA pol activity used to make complementary DNA resulting in RNA:DNA hybrid iii) Ribonuclease activity degrades the RNA strand iv) DNA dependent DNA pol makes copy of the DNA strand resulting in dsDNA. Problems in Viral Replication Which of the following would best explain the high mutation rate of RNA viruses and their ability to evolve rapidly? A. They rely on host RNA polymerase B. They rely on DNA Polymerase III which lacks proofreading C. They rely on DNA Polymerase I which lacks proofreading D. They rely on viral polymerases which have high error rates E. Ribosomes have high error rates when translating RNA. Problems in Viral Replication viruses need RNA Replicase (RNA dependent RNA polymerase) viruses need Reverse Transcriptase (RNA dependent DNA poly While a few animal viruses have DNA genomes, most animal viruses have RNA genomes. Which of the following would best explain why DNA viruses are rare in animals, but not rare in Bacteria? A. Eukarya compartmentalize transcription and DNA replication to the nucleus B. Eukarya degrade foreign DNA in the cytoplasm C. Eukarya translate mRNA in the cytoplasm D. Eukarya lack DNA polymerase E. Mitosis prevents replication of viral DNA. Problems in Viral Replication ircular ssDNA (+) Virus: X 174. Problems in Viral Replication near ssRNA (-) Virus: Rhabdovirus -Viral Replicase: RNA dependent RNA pol -Protein primer: Protein acts as primer for replicase. Problems in Viral Replication he ‘end problem’ of replicating linear genomes Solutions: Terminal redundancy (e.g. T4 Phage) Make circular DNA (e.g. Lambda) Terminal repeats (e.g. HIV, Telomeres) Terminal proteins as primers Summary Points: All viruses must produce mRNA in order to make viral proteins Some viruses (temperate phage, retroviruses) integrate into host DNA Linear viruses need a mechanism for replicating their ends RNA viruses have high mutation rates because their viral polymerases (replicase or reverse transcriptase)