Characterization of HIV-1 uncoating in human microglial cell lines.

Virology Journal
Zachary IngramAmy E Hulme

Abstract

After viral fusion with the cell membrane, the conical capsid of HIV-1 disassembles by a process called uncoating. Previously we have utilized the CsA washout assay, in which TRIM-CypA mediated restriction of viral replication is used to detect the state of the viral capsid, to study the kinetics of HIV-1 uncoating in owl monkey kidney (OMK) and HeLa cells. Here we have extended this analysis to the human microglial cell lines CHME3 and C20 to characterize uncoating in a cell type that is a natural target of HIV infection. The CsA washout was used to characterize uncoating of wildtype and capsid mutant viruses in CHME3 and C20 cells. Viral fusion assays and nevirapine addition assays were performed to relate the kinetics of viral fusion and reverse transcription to uncoating. We found that uncoating initiated within the first hour after viral fusion and was facilitated by reverse transcription in CHME3 and C20 cells. The capsid mutation A92E did not significantly alter uncoating kinetics. Viruses with capsid mutations N74D and E45A decreased the rate of uncoating in CHME3 cells, but did not alter reverse transcription. Interestingly, the second site suppressor capsid mutation R132T was able to rescue the uncoating kinetics of t...Continue Reading

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Citations

Sep 9, 2020·The EMBO Journal·Eveline Santos da SilvaMojgan H Naghavi
Feb 10, 2021·Viruses·Morganne Wilbourne, Peijun Zhang
Jun 3, 2021·Viruses·Maryam TabasiZeger Debyser

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Methods Mentioned

BETA
fluorescence microscopy
transfection
flow cytometry
atomic force microscopy
fluorescence imaging

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