Nonenveloped viruses must navigate a number of host cell membrane barriers during entry and after replication for release. Simian Virus 40 (SV40) pirates the endocytic pathway of the host cell for transport from the cell surface until it reaches the endoplasmic reticulum (ER). Within the ER lumen the capsid is thought to be disassembled releasing the viral genome which then must be transported across the ER and nuclear membrane barriers to initiate viral replication. Following replication the SV40 viral progeny must cross the nuclear plasma membrane without becoming membrane encapsulated. To overcome these obstacles during release, viruses such as SV40 utilize a lytic infectious cycle that results in the permeabilization of the host cell membranes through an unknown mechanism. The research presented here utilized numerous techniques to investigate the roles of the minor structural proteins in the SV40 life cycle. VP2 and VP3 were both shown to perform essential functions with VP2 contributing to cell binding while VP3 functioned downstream of this event. Both VP3 and VP2 showed the ability to post-translationally integrate into the ER membrane where VP3 attained a multi-membrane spanning topology. These findings combined with the propensity of VP2 and VP3 to oligomerize and permeabilize bacterial membranes provide the premise for the following hypothesis. We propose that upon disassembly in the ER lumen these minor structural oligomerize and integrate into the ER membrane where they act as a viroporin to aid in the transfer of the viral genome across the ER membrane. The SV40 mutant lacking both the VP2 and VP3 initiation codons revealed that in addition to VP3, another viral protein was synthesized from the VP2 reading frame. This protein we termed VP4 is expressed at the onset of lysis, it is not sequestered by VP1 and its removal inhibited the lytic property of VP3 towards E. coli and caused a significant 2 day delay in cell lysis during infection. In summary, my work provides new insight into how nonenveloped viruses have evolved to traverse the host cell membrane barriers during entry, when they must remain intact, and during release when they become dispensible.
| Identifer | oai:union.ndltd.org:UMASS/oai:scholarworks.umass.edu:dissertations-4558 |
| Date | 01 January 2007 |
| Creators | Daniels, Robert D |
| Publisher | ScholarWorks@UMass Amherst |
| Source Sets | University of Massachusetts, Amherst |
| Language | English |
| Detected Language | English |
| Type | text |
| Source | Doctoral Dissertations Available from Proquest |
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