Identification of a Structural Element in HIV-1 Gag Required for Virus Particle Assembly and Maturation

ABSTRACT Late in the HIV-1 replication cycle, the viral structural protein Gag is targeted to virus assembly sites at the plasma membrane of infected cells. The capsid (CA) domain of Gag plays a critical role in the formation of the hexameric Gag lattice in the immature virion, and, during particle...

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Autores principales:	Mariia Novikova, Lucas J. Adams, Juan Fontana, Anna T. Gres, Muthukumar Balasubramaniam, Dennis C. Winkler, Sagar B. Kudchodkar, Ferri Soheilian, Stefan G. Sarafianos, Alasdair C. Steven, Eric O. Freed
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2018
Materias:	Cryo-EM HIV-1 assembly capsid maturation virus replication Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/d1bc164630d541f6abb520b9b82a3d58
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spelling	oai:doaj.org-article:d1bc164630d541f6abb520b9b82a3d582021-11-15T15:58:20ZIdentification of a Structural Element in HIV-1 Gag Required for Virus Particle Assembly and Maturation10.1128/mBio.01567-182150-7511https://doaj.org/article/d1bc164630d541f6abb520b9b82a3d582018-11-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.01567-18https://doaj.org/toc/2150-7511ABSTRACT Late in the HIV-1 replication cycle, the viral structural protein Gag is targeted to virus assembly sites at the plasma membrane of infected cells. The capsid (CA) domain of Gag plays a critical role in the formation of the hexameric Gag lattice in the immature virion, and, during particle release, CA is cleaved from the Gag precursor by the viral protease and forms the conical core of the mature virion. A highly conserved Pro-Pro-Ile-Pro (PPIP) motif (CA residues 122 to 125) [PPIP(122–125)] in a loop connecting CA helices 6 and 7 resides at a 3-fold axis formed by neighboring hexamers in the immature Gag lattice. In this study, we characterized the role of this PPIP(122–125) loop in HIV-1 assembly and maturation. While mutations P123A and P125A were relatively well tolerated, mutation of P122 and I124 significantly impaired virus release, caused Gag processing defects, and abolished infectivity. X-ray crystallography indicated that the P122A and I124A mutations induce subtle changes in the structure of the mature CA lattice which were permissive for in vitro assembly of CA tubes. Transmission electron microscopy and cryo-electron tomography demonstrated that the P122A and I124A mutations induce severe structural defects in the immature Gag lattice and abrogate conical core formation. Propagation of the P122A and I124A mutants in T-cell lines led to the selection of compensatory mutations within CA. Our findings demonstrate that the CA PPIP(122–125) loop comprises a structural element critical for the formation of the immature Gag lattice. IMPORTANCE Capsid (CA) plays multiple roles in the HIV-1 replication cycle. CA-CA domain interactions are responsible for multimerization of the Gag polyprotein at virus assembly sites, and in the mature virion, CA monomers assemble into a conical core that encapsidates the viral RNA genome. Multiple CA regions that contribute to the assembly and release of HIV-1 particles have been mapped and investigated. Here, we identified and characterized a Pro-rich loop in CA that is important for the formation of the immature Gag lattice. Changes in this region disrupt viral production and abrogate the formation of infectious, mature virions. Propagation of the mutants in culture led to the selection of second-site compensatory mutations within CA. These results expand our knowledge of the assembly and maturation steps in the viral replication cycle and may be relevant for development of antiviral drugs targeting CA.Mariia NovikovaLucas J. AdamsJuan FontanaAnna T. GresMuthukumar BalasubramaniamDennis C. WinklerSagar B. KudchodkarFerri SoheilianStefan G. SarafianosAlasdair C. StevenEric O. FreedAmerican Society for MicrobiologyarticleCryo-EMHIV-1assemblycapsidmaturationvirus replicationMicrobiologyQR1-502ENmBio, Vol 9, Iss 5 (2018)
institution	DOAJ
collection	DOAJ
language	EN
topic	Cryo-EM HIV-1 assembly capsid maturation virus replication Microbiology QR1-502
spellingShingle	Cryo-EM HIV-1 assembly capsid maturation virus replication Microbiology QR1-502 Mariia Novikova Lucas J. Adams Juan Fontana Anna T. Gres Muthukumar Balasubramaniam Dennis C. Winkler Sagar B. Kudchodkar Ferri Soheilian Stefan G. Sarafianos Alasdair C. Steven Eric O. Freed Identification of a Structural Element in HIV-1 Gag Required for Virus Particle Assembly and Maturation
description	ABSTRACT Late in the HIV-1 replication cycle, the viral structural protein Gag is targeted to virus assembly sites at the plasma membrane of infected cells. The capsid (CA) domain of Gag plays a critical role in the formation of the hexameric Gag lattice in the immature virion, and, during particle release, CA is cleaved from the Gag precursor by the viral protease and forms the conical core of the mature virion. A highly conserved Pro-Pro-Ile-Pro (PPIP) motif (CA residues 122 to 125) [PPIP(122–125)] in a loop connecting CA helices 6 and 7 resides at a 3-fold axis formed by neighboring hexamers in the immature Gag lattice. In this study, we characterized the role of this PPIP(122–125) loop in HIV-1 assembly and maturation. While mutations P123A and P125A were relatively well tolerated, mutation of P122 and I124 significantly impaired virus release, caused Gag processing defects, and abolished infectivity. X-ray crystallography indicated that the P122A and I124A mutations induce subtle changes in the structure of the mature CA lattice which were permissive for in vitro assembly of CA tubes. Transmission electron microscopy and cryo-electron tomography demonstrated that the P122A and I124A mutations induce severe structural defects in the immature Gag lattice and abrogate conical core formation. Propagation of the P122A and I124A mutants in T-cell lines led to the selection of compensatory mutations within CA. Our findings demonstrate that the CA PPIP(122–125) loop comprises a structural element critical for the formation of the immature Gag lattice. IMPORTANCE Capsid (CA) plays multiple roles in the HIV-1 replication cycle. CA-CA domain interactions are responsible for multimerization of the Gag polyprotein at virus assembly sites, and in the mature virion, CA monomers assemble into a conical core that encapsidates the viral RNA genome. Multiple CA regions that contribute to the assembly and release of HIV-1 particles have been mapped and investigated. Here, we identified and characterized a Pro-rich loop in CA that is important for the formation of the immature Gag lattice. Changes in this region disrupt viral production and abrogate the formation of infectious, mature virions. Propagation of the mutants in culture led to the selection of second-site compensatory mutations within CA. These results expand our knowledge of the assembly and maturation steps in the viral replication cycle and may be relevant for development of antiviral drugs targeting CA.
format	article
author	Mariia Novikova Lucas J. Adams Juan Fontana Anna T. Gres Muthukumar Balasubramaniam Dennis C. Winkler Sagar B. Kudchodkar Ferri Soheilian Stefan G. Sarafianos Alasdair C. Steven Eric O. Freed
author_facet	Mariia Novikova Lucas J. Adams Juan Fontana Anna T. Gres Muthukumar Balasubramaniam Dennis C. Winkler Sagar B. Kudchodkar Ferri Soheilian Stefan G. Sarafianos Alasdair C. Steven Eric O. Freed
author_sort	Mariia Novikova
title	Identification of a Structural Element in HIV-1 Gag Required for Virus Particle Assembly and Maturation
title_short	Identification of a Structural Element in HIV-1 Gag Required for Virus Particle Assembly and Maturation
title_full	Identification of a Structural Element in HIV-1 Gag Required for Virus Particle Assembly and Maturation
title_fullStr	Identification of a Structural Element in HIV-1 Gag Required for Virus Particle Assembly and Maturation
title_full_unstemmed	Identification of a Structural Element in HIV-1 Gag Required for Virus Particle Assembly and Maturation
title_sort	identification of a structural element in hiv-1 gag required for virus particle assembly and maturation
publisher	American Society for Microbiology
publishDate	2018
url	https://doaj.org/article/d1bc164630d541f6abb520b9b82a3d58
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Identification of a Structural Element in HIV-1 Gag Required for Virus Particle Assembly and Maturation

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