The Host Cell Metabolite Inositol Hexakisphosphate Promotes Efficient Endogenous HIV-1 Reverse Transcription by Stabilizing the Viral Capsid

ABSTRACT A defining activity of retroviruses is reverse transcription, the process by which the viral genomic RNA is converted into the double-stranded DNA required for virus replication. Reverse transcriptase (RT), the viral enzyme responsible for this process, was identified in 1970 by assaying pe...

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Autores principales:	Jordan Jennings, Jiong Shi, Janani Varadarajan, Parker J. Jamieson, Christopher Aiken
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2020
Materias:	HIV-1 IP6 capsid reverse transcription uncoating Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/9f4588bdb95c474798f388647168f867
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spelling	oai:doaj.org-article:9f4588bdb95c474798f388647168f8672021-11-15T15:55:43ZThe Host Cell Metabolite Inositol Hexakisphosphate Promotes Efficient Endogenous HIV-1 Reverse Transcription by Stabilizing the Viral Capsid10.1128/mBio.02820-202150-7511https://doaj.org/article/9f4588bdb95c474798f388647168f8672020-12-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.02820-20https://doaj.org/toc/2150-7511ABSTRACT A defining activity of retroviruses is reverse transcription, the process by which the viral genomic RNA is converted into the double-stranded DNA required for virus replication. Reverse transcriptase (RT), the viral enzyme responsible for this process, was identified in 1970 by assaying permeabilized retrovirus particles for DNA synthesis in vitro. Such reactions are inefficient, with only a small fraction of viral genomes being converted to full-length double-stranded DNA molecules, possibly owing to disruption of the structure of the viral core. Here, we show that reverse transcription in purified HIV-1 cores is enhanced by the addition of the capsid-binding host cell metabolite inositol hexakisphosphate (IP6). IP6 potently enhanced full-length minus-strand synthesis, as did hexacarboxybenzene (HCB), which also stabilizes the HIV-1 capsid. Both IP6 and HCB stabilized the association of the viral CA and RT proteins with HIV-1 cores. In contrast to the wild type, cores isolated from mutant HIV-1 particles containing intrinsically hyperstable capsids exhibited relatively efficient reverse transcription in the absence of IP6, further indicating that the compound promotes reverse transcription by stabilizing the viral capsid. We also observed that the capsid-destabilizing antiviral compound PF74 inhibited endogenous reverse transcription with a potency that mirrors its ability to inhibit reverse transcription during infection. Our results show that the stabilization of the HIV-1 capsid permits efficient reverse transcription in HIV-1 cores, providing a sensitive experimental system for analyzing the functions of viral and host cell molecules and the role of capsid disassembly (uncoating) in the process. IMPORTANCE HIV-1 infection requires reverse transcription of the viral genome. While much is known about the biochemistry of reverse transcription from simplified biochemical reactions, reverse transcription during infection takes place within a viral core. However, endogenous reverse transcription reactions using permeabilized HIV-1 virions or purified viral cores have been inefficient. Using viral cores purified from infectious HIV-1 particles, we show that efficient reverse transcription is achieved in vitro by addition of the capsid-stabilizing metabolite inositol hexakisphosphate. The enhancement of reverse transcription was linked to the capsid-stabilizing effect of the compound, consistent with the known requirement for an intact or semi-intact viral capsid for HIV-1 infection. Our results establish a biologically relevant system for dissecting the function of the viral capsid and its disassembly during reverse transcription. The system should also prove useful for mechanistic studies of capsid-targeting antiviral drugs.Jordan JenningsJiong ShiJanani VaradarajanParker J. JamiesonChristopher AikenAmerican Society for MicrobiologyarticleHIV-1IP6capsidreverse transcriptionuncoatingMicrobiologyQR1-502ENmBio, Vol 11, Iss 6 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	HIV-1 IP6 capsid reverse transcription uncoating Microbiology QR1-502
spellingShingle	HIV-1 IP6 capsid reverse transcription uncoating Microbiology QR1-502 Jordan Jennings Jiong Shi Janani Varadarajan Parker J. Jamieson Christopher Aiken The Host Cell Metabolite Inositol Hexakisphosphate Promotes Efficient Endogenous HIV-1 Reverse Transcription by Stabilizing the Viral Capsid
description	ABSTRACT A defining activity of retroviruses is reverse transcription, the process by which the viral genomic RNA is converted into the double-stranded DNA required for virus replication. Reverse transcriptase (RT), the viral enzyme responsible for this process, was identified in 1970 by assaying permeabilized retrovirus particles for DNA synthesis in vitro. Such reactions are inefficient, with only a small fraction of viral genomes being converted to full-length double-stranded DNA molecules, possibly owing to disruption of the structure of the viral core. Here, we show that reverse transcription in purified HIV-1 cores is enhanced by the addition of the capsid-binding host cell metabolite inositol hexakisphosphate (IP6). IP6 potently enhanced full-length minus-strand synthesis, as did hexacarboxybenzene (HCB), which also stabilizes the HIV-1 capsid. Both IP6 and HCB stabilized the association of the viral CA and RT proteins with HIV-1 cores. In contrast to the wild type, cores isolated from mutant HIV-1 particles containing intrinsically hyperstable capsids exhibited relatively efficient reverse transcription in the absence of IP6, further indicating that the compound promotes reverse transcription by stabilizing the viral capsid. We also observed that the capsid-destabilizing antiviral compound PF74 inhibited endogenous reverse transcription with a potency that mirrors its ability to inhibit reverse transcription during infection. Our results show that the stabilization of the HIV-1 capsid permits efficient reverse transcription in HIV-1 cores, providing a sensitive experimental system for analyzing the functions of viral and host cell molecules and the role of capsid disassembly (uncoating) in the process. IMPORTANCE HIV-1 infection requires reverse transcription of the viral genome. While much is known about the biochemistry of reverse transcription from simplified biochemical reactions, reverse transcription during infection takes place within a viral core. However, endogenous reverse transcription reactions using permeabilized HIV-1 virions or purified viral cores have been inefficient. Using viral cores purified from infectious HIV-1 particles, we show that efficient reverse transcription is achieved in vitro by addition of the capsid-stabilizing metabolite inositol hexakisphosphate. The enhancement of reverse transcription was linked to the capsid-stabilizing effect of the compound, consistent with the known requirement for an intact or semi-intact viral capsid for HIV-1 infection. Our results establish a biologically relevant system for dissecting the function of the viral capsid and its disassembly during reverse transcription. The system should also prove useful for mechanistic studies of capsid-targeting antiviral drugs.
format	article
author	Jordan Jennings Jiong Shi Janani Varadarajan Parker J. Jamieson Christopher Aiken
author_facet	Jordan Jennings Jiong Shi Janani Varadarajan Parker J. Jamieson Christopher Aiken
author_sort	Jordan Jennings
title	The Host Cell Metabolite Inositol Hexakisphosphate Promotes Efficient Endogenous HIV-1 Reverse Transcription by Stabilizing the Viral Capsid
title_short	The Host Cell Metabolite Inositol Hexakisphosphate Promotes Efficient Endogenous HIV-1 Reverse Transcription by Stabilizing the Viral Capsid
title_full	The Host Cell Metabolite Inositol Hexakisphosphate Promotes Efficient Endogenous HIV-1 Reverse Transcription by Stabilizing the Viral Capsid
title_fullStr	The Host Cell Metabolite Inositol Hexakisphosphate Promotes Efficient Endogenous HIV-1 Reverse Transcription by Stabilizing the Viral Capsid
title_full_unstemmed	The Host Cell Metabolite Inositol Hexakisphosphate Promotes Efficient Endogenous HIV-1 Reverse Transcription by Stabilizing the Viral Capsid
title_sort	host cell metabolite inositol hexakisphosphate promotes efficient endogenous hiv-1 reverse transcription by stabilizing the viral capsid
publisher	American Society for Microbiology
publishDate	2020
url	https://doaj.org/article/9f4588bdb95c474798f388647168f867
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The Host Cell Metabolite Inositol Hexakisphosphate Promotes Efficient Endogenous HIV-1 Reverse Transcription by Stabilizing the Viral Capsid

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