Dodding, Mark Peter; (2007) Interactions of retroviral capsid proteins with restriction factors. Doctoral thesis , UCL (University College London).

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Abstract

It is becoming increasingly clear that mammalian cells can express a variety of factors that limit retroviral replication. One class of these factors blocks replication at a stage post-entry but prior to nuclear import and integration of the viral genome. Members of this family arc known as restriction factors. The best characterised member is the mouse gene h'vl which determines susceptibility to infection by MLV. I-'vl has two alleles known as Fvl" and Fv/' whose restriction characteristics determine the host range of different MLV strains. It has become apparent that the phenomenon of restriction is not limited to MLV and murine cells. HIV-1, as well as MLV is restricted at a post-entry step in a number of primate species including rhesus macaques. African green monkeys and owl monkeys. The Thm5 gene in these species is the main determinant of both HIV-1 and MLV restriction. Viral determinants of this restriction map to the virus capsid protein, however despite strong genetic ev idence, no direct interaction has been shown between capsid and restriction factor using a variety of biochemical approaches. This project began by aiming to understand why this might be the case. Experiments using a series of mutants defective for proper processing and assembly of capsid yielded evidence suggesting that the restriction factor binding site is in fact formed only when capsid is in its polymeric state in a mature virus, thus explaining why conventional approaches had failed to detect any interaction. This also provided insight into the mode of action of restriction factors, suggesting that they have evolved to target assembled mature virus entering the cytoplasm. In order to better understand the context of the capsid/restriction factor interaction, cell biology studies using live-cell microscopy were initiated. These revealed that Trim5u is found in association with the microtubule network in highly motile cytoplasmic bodies, suggesting the possibility that the incoming virus might interaction w ith Trim5u in this context. furthermore, structural studies aimed at obtaining a better understanding of the structure of the capsid core have suggested new interactions w hich may be important for its formation, stability and consequent infectivity of the virus. These offer a potential new target for anti-retroviral therapies.

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