HIV-1 Gag co-opts a cellular complex containing DDX6, a helicase that facilitates capsid assembly

Reed, Jonathan C.; Molter, Britta; Geary, Clair D.; McNevin, John; McElrath, M. Juliana; Giri, Samina; Klein, Kevin; Lingappa, Jaisri R.

doi:10.1083/jcb.201111012

Cited by 78 publications

(221 citation statements)

References 84 publications

(160 reference statements)

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“…These complexes are true assembly intermediates, since they appear in a sequential progression in pulse-chase analyses, with newly synthesized Gag first forming the ϳ10S complex, followed by the ϳ80S intermediate, ϳ150S intermediate, and ϳ500S intermediate, before culminating in the ϳ750S completely assembled HIV-1 immature capsid (40,41). The order of intermediates in the assembly pathway initially indicated by pulse-chase experiments has been confirmed through the analysis of assembly-defective mutants (40,(42)(43)(44). In these studies, cells expressing an assembly-defective Gag mutant accumulate only those intermediates that precede the step at which assembly is blocked.…”

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confidence: 81%

“…In these studies, cells expressing an assembly-defective Gag mutant accumulate only those intermediates that precede the step at which assembly is blocked. The ϳ80S, ϳ150S, and ϳ500S assembly intermediates (termed the high-molecular-mass intermediates) contain Gag as well as cellular proteins derived from a host precursor complex, at least two of which-the ATP-binding cassette protein E1 (ABCE1) and the DEAD box RNA helicase DDX6 -facilitate immature capsid assembly (40,42,43,45,46). Formation of the ϳ80S intermediate, in which Gag first associates with ABCE1 and DDX6, occurs when Gag co-opts the cellular precursor complex that contains these and other host proteins (43).…”

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confidence: 99%

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A Temporospatial Map That Defines Specific Steps at Which Critical Surfaces in the Gag MA and CA Domains Act during Immature HIV-1 Capsid Assembly in Cells

Robinson

Reed

Geary

et al. 2014

J Virol

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147

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During HIV-1 assembly, Gag polypeptides target to the plasma membrane, where they multimerize to form immature capsids that undergo budding and maturation. Previous mutational analyses identified residues within the Gag matrix (MA) and capsid (CA) domains that are required for immature capsid assembly, and structural studies showed that these residues are clustered on four exposed surfaces in Gag. Exactly when and where the three critical surfaces in CA function during assembly are not known. Here, we analyzed how mutations in these four critical surfaces affect the formation and stability of assembly intermediates in cells expressing the HIV-1 provirus. The resulting temporospatial map reveals that critical MA residues act during membrane targeting, residues in the C-terminal CA subdomain (CA-CTD) dimer interface are needed for the stability of the first membrane-bound assembly intermediate, CA-CTD base residues are necessary for progression past the first membrane-bound intermediate, and residues in the N-terminal CA subdomain (CA-NTD) stabilize the last membrane-bound intermediate. Importantly, we found that all four critical surfaces act while Gag is associated with the cellular facilitators of assembly ABCE1 and DDX6. When correlated with existing structural data, our findings suggest the following model: Gag dimerizes via the CA-CTD dimer interface just before or during membrane targeting, individual CA-CTD hexamers form soon after membrane targeting, and the CA-NTD hexameric lattice forms just prior to capsid release. This model adds an important new dimension to current structural models by proposing the potential order in which key contacts within the immature capsid lattice are made during assembly in cells. IMPORTANCEWhile much is known about the structure of the completed HIV-1 immature capsid and domains of its component Gag proteins, less is known about the sequence of events leading to formation of the HIV-1 immature capsid. Here we used biochemical and ultrastructural analyses to generate a temporospatial map showing the precise order in which four critical surfaces in Gag act during immature capsid formation in provirus-expressing cells. Because three of these surfaces make important contacts in the hexameric lattices that are found in the completed immature capsid, these data allow us to propose a model for the sequence of events leading to formation of the hexameric lattices. By providing a dynamic view of when and where critical Gag-Gag contacts form during the assembly process and how those contacts function in the nascent capsid, our study provides novel insights into how an immature capsid is built in infected cells.

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confidence: 81%

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confidence: 99%

A Temporospatial Map That Defines Specific Steps at Which Critical Surfaces in the Gag MA and CA Domains Act during Immature HIV-1 Capsid Assembly in Cells

Robinson

Reed

Geary

et al. 2014

J Virol

Self Cite

147

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“…In this regard, it is worthwhile to note the interconnections between translation and RNA-processing proteins. In particular, the destinations of viral and cellular RNAs include actively translating ribosomes; stress granules (SGs), where mRNAs on stalled preinitiation complexes are stored or sorted; and processing bodies (PBs), involved in RNA silencing and decay (22,(90)(91)(92)(93)(94)(95). Previously, HIV-1 Gag proteins were found to associate with elongation factor 1 alpha, via RNA bridges to MA and NC, and this interaction was proposed to induce a shift from Gag protein translation to vRNA encapsidation (26).…”

Section: Discussionmentioning

confidence: 99%

Analysis of HIV-1 Gag Protein Interactions via Biotin Ligase Tagging

Ritchie

Cylinder

Platt

et al. 2015

J Virol

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We have examined the interactions of wild-type (WT) and matrix protein-deleted (⌬MA) HIV-1 precursor Gag (PrGag) proteins in virus-producing cells using a biotin ligase-tagging approach. To do so, WT and ⌬MA PrGag proteins were tagged with the Escherichia coli promiscuous biotin ligase (BirA*), expressed in cells, and examined. Localization patterns of PrGag proteins and biotinylated proteins overlapped, consistent with observations that BirA*-tagged proteins biotinylate neighbor proteins that are in close proximity. Results indicate that BirA*-tagged PrGag proteins biotinylated themselves as well as WT PrGag proteins in trans. Previous data have shown that the HIV-1 Envelope (Env) protein requires an interaction with MA for assembly into virions. Unexpectedly, ⌬MA proteins biotinylated Env, whereas WT BirA*-tagged proteins did not, suggesting that the presence of MA made Env inaccessible to biotinylation. We also identified over 50 cellular proteins that were biotinylated by BirA*-tagged PrGag proteins. These included membrane proteins, cytoskeleton-associated proteins, nuclear transport factors, lipid metabolism regulators, translation factors, and RNA-processing proteins. The identification of these biotinylated proteins offers new insights into HIV-1 Gag protein trafficking and activities and provides new potential targets for antiviral interference.

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“…However, studies disagree about the positive contributions of PB factors to retrovirus infection. DEAD box helicase DDX6 (ScDhh1) is ascribed a positive role in the intracellular packaging of prototypic foamy virus RNA, but studies differ as to roles in HIV-1 core assembly (224,225). Overall these studies suggest that individual components of ribonucleoprotein granules, including PB and SG, interact with assembling retrovirus RNA and proteins, but assembly is not associated with these granules per se.…”

Section: Figurementioning

confidence: 82%

Ty3, a Position-specific Retrotransposon in Budding Yeast

Sandmeyer

Patterson²,

Bilanchone³

2015

Microbiol Spectr

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Long terminal repeat (LTR) retrotransposons constitute significant fractions of many eukaryotic genomes. Two ancient families are Ty1/Copia (Pseudoviridae) and Ty3/Gypsy (Metaviridae). The Ty3/Gypsy family probably gave rise to retroviruses based on the domain order, similarity of sequences, and the envelopes encoded by some members. The Ty3 element of Saccharomyces cerevisiae is one of the most completely characterized elements at the molecular level. Ty3 is induced in mating cells by pheromone stimulation of the mitogen-activated protein kinase pathway as cells accumulate in G1. The two Ty3 open reading frames are translated into Gag3 and Gag3-Pol3 polyprotein precursors. In haploid mating cells Gag3 and Gag3-Pol3 are assembled together with Ty3 genomic RNA into immature virus-like particles in cellular foci containing RNA processing body proteins. Virus-like particle Gag3 is then processed by Ty3 protease into capsid, spacer, and nucleocapsid, and Gag3-Pol3 into those proteins and additionally, protease, reverse transcriptase, and integrase. After haploid cells mate and become diploid, genomic RNA is reverse transcribed into cDNA. Ty3 integration complexes interact with components of the RNA polymerase III transcription complex resulting in Ty3 integration precisely at the transcription start site. Ty3 activation during mating enables proliferation of Ty3 between genomes and has intriguing parallels with metazoan retrotransposon activation in germ cell lineages. Identification of nuclear pore, DNA replication, transcription, and repair host factors that affect retrotransposition has provided insights into how hosts and retrotransposons interact to balance genome stability and plasticity.

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HIV-1 Gag co-opts a cellular complex containing DDX6, a helicase that facilitates capsid assembly

Abstract: The RNA helicase DDX6 promotes HIV-1 assembly in a co-opted cellular complex containing P body proteins and ABCE1.

Cited by 78 publications

References 84 publications

A Temporospatial Map That Defines Specific Steps at Which Critical Surfaces in the Gag MA and CA Domains Act during Immature HIV-1 Capsid Assembly in Cells

A Temporospatial Map That Defines Specific Steps at Which Critical Surfaces in the Gag MA and CA Domains Act during Immature HIV-1 Capsid Assembly in Cells

Analysis of HIV-1 Gag Protein Interactions via Biotin Ligase Tagging

Ty3, a Position-specific Retrotransposon in Budding Yeast

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