Miles, Richard Joseph; (2018) Investigating the anti-HIV-1 restriction factor MxB. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

The interferon inducible dynamin-like GTPase MxB restricts HIV-1 infection in various cell types at the stage of viral nuclear entry. Recent evidence suggests that capsid is the mediator of viral susceptibility to MxB, possibly through binding of MxB to the viral core inhibiting uncoating. We have shown that MxB restriction is independent of the conserved nuclear entry pathway used by HIV-1 supporting the model of capsid binding. Knockdown of cyclophilin A or treatment with cyclosporine A fully rescued wild-type HIV-1 from MxB restriction. This suggests that MxB restriction requires the presence of CypA and its ability to recruit to the HIV-1 capsid to block infection. In addition, there is varying sensitivity to MxB restriction of different capsid isolates within both M- and O- group subtypes. As well as blocking viral infection, many restriction factors also act as molecular tripwires - initiating innate immune signaling pathways upon viral engagement. Here we demonstrate that MxB, but not MxA, is capable of activating both NFκB and AP-1 promoters when overexpressed in 293T cells in a dose-dependent manner and independent of any other known activation signal. While a functional GTPase domain is not required for restriction, GTP binding but not hydrolysis is absolutely required for NFκB activation. Furthermore, it is necessary to communicate conformational changes induced upon GTP binding to robustly activate NFκB signaling. Finally, we have developed techniques to study the spatial localisation of the HIV-1 provirus within the nucleus by DNA FISH. We can also measure transcriptional output at a single-cell level by RNA FISH and correlate this to proviral nuclear position. We discovered that HIV-1 displays some nuclear spatial preference to integration and that this localisation may impact on the transcriptional output of the integrated provirus.

Type:	Thesis (Doctoral)
Qualification:	Ph.D
Title:	Investigating the anti-HIV-1 restriction factor MxB
Event:	UCL
Language:	English
UCL classification:	UCL UCL > Provost and Vice Provost Offices UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Medical Sciences > Div of Infection and Immunity
URI:	https://discovery.ucl.ac.uk/id/eprint/10042080

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