Emergence of Compensatory Mutations Reveals the Importance of Electrostatic Interactions between HIV-1 Integrase and Genomic RNA

Mugisha, Christian Shema; Dinh, Tung; Kumar, Abhishek; Tenneti, Kasyap; Eschbach, Jenna E.; Davis, Keanu; Gifford, Robert J.; Kvaratskhelia, Mamuka; Kutluay, Sebla B.

doi:10.1128/mbio.00431-22

Cited by 10 publications

(11 citation statements)

References 105 publications

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“…Prior studies suggested that the formation of a core IN tetramer is relevant for the proper execution of two distinct functions mediated by IN, integration 9 and RNA binding 24,25,35 . We set out to determine the structure of the WT IN tetramer in apo form without nucleic acids bound.…”

Section: Resultsmentioning

confidence: 99%

“…The arrangement of the tetrameric IN assembly has important implications for a working model of its binding to RNA. The four CTDs, which are crucial for RNA binding 24,25,35 , zigzag along the interface of two IN dimers ( Figure 6a ). These CTDs provide a highly positively charged surface stretching over 80 Å, which could effectively bind RNA ( Figure 6b ).…”

Section: Discussionmentioning

confidence: 99%

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Oligomeric HIV-1 Integrase Structures Reveal Functional Plasticity for Intasome Assembly and RNA Binding

Jing,

Shan,

Dinh

et al. 2024

Preprint

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Integrase (IN) performs dual essential roles during HIV-1 replication. During ingress, IN functions within an oligomeric intasome assembly to catalyze viral DNA integration into host chromatin. During late stages of infection, tetrameric IN binds viral RNA and orchestrates the condensation of ribonucleoprotein complexes into the capsid core. The molecular architectures of IN assemblies that mediate these distinct events remain incompletely characterized. Furthermore, the tetramer is an important antiviral target for allosteric IN inhibitors. Here, we determined cryo-EM structures of wildtype HIV-1 IN tetramers and intasome hexadecamers. Our structures unveil a remarkable plasticity that leverages IN C-terminal domains and abutting linkers to assemble functionally distinct oligomeric forms. Alteration of a newly recognized conserved interface revealed that both IN functions track with tetramerization in vitro and during HIV-1 infection. Collectively, our findings reveal how IN plasticity orchestrates its diverse molecular functions and provide a working model for IN-viral RNA binding and atomic blueprints for allosteric IN inhibitor development.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Oligomeric HIV-1 Integrase Structures Reveal Functional Plasticity for Intasome Assembly and RNA Binding

Jing,

Shan,

Dinh

et al. 2024

Preprint

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“…Thus, the GagPol-containing IN:M50I/V151I may differentially regulate the ESCRT system and suppress virus release. Recent studies highlight accessory functions of matured IN (52–54), IN stimulates RT (52), IN interacts with RNA(54–56), acetylation of CTD of IN is associated with defects in proviral transcription (55). We now demonstrate a new potential role of IN domain in GagPol polyprotein.…”

Section: Discussionmentioning

confidence: 99%

The C-terminal domain of RNase H and the C-terminus amino acid residue regulate virus release and autoprocessing of a defective HIV-1 possessing M50I and V151I changes in integrase

Imamichi

Chen

Hao

et al. 2022

Preprint

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Previously, we reported that an HIV-1 variant containing Met-to-Ile change at codon 50 and Val-to-Ile mutation at codon 151 of integrase (IN), HIV(IN:M50I/V151I), was an impaired virus. Despite the mutations being in IN, the virus release was significantly suppressed (p < 0.0001) and the initiation of autoprocessing was inhibited; the mechanism of the defect remains unknown. In the current study, we attempted to identify the critical domains or amino acid (aa) residue(s) that promote defects in HIV(IN:M50I/V151I), using a series of variants, including truncated or aa-substituted RNase H (RH) or IN. The results demonstrated that virus release and the initiation of autoprocessing were regulated by the C-terminal domains (CTDs) of RH and IN. Further studies illustrated that Asp at codon 109 of RH CTD and Asp at the C terminus of IN induces the defect. This result indicated that the CTD of RH and IN in GagPol and particular aa positions in RH and IN regulated the virus release and the initiation of autoprocessing, and these sites could be potential targets for the developing new therapies.

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“…The quantity of HIV-1 RT products and 2-LTR circles was measured by qPCR using primers specific for early and late RT products (see above table). Extracted DNA was analyzed for accumulation of RT products, 2-LTR circles, and integration as described before ( 97 , 98 ).…”

Section: Methodsmentioning

confidence: 99%

HIV-1 capsid stability and reverse transcription are finely balanced to minimize sensing of reverse transcription products via the cGAS-STING pathway

Eschbach,

Puray-Chavez,

Mohammed

et al. 2024

mBio

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A critical determinant for early post-entry events, the HIV-1 capsid (CA) protein forms the conical core when it rearranges around the dimeric RNA genome and associated viral proteins. Although mutations in CA have been reported to alter innate immune sensing of HIV-1, a direct link between core stability and sensing of HIV-1 nucleic acids has not been established. Herein, we assessed how manipulating the stability of the CA lattice through chemical and genetic approaches affects innate immune recognition of HIV-1. We found that destabilization of the CA lattice resulted in potent sensing of reverse transcription products when destabilization per se does not completely block reverse transcription. Surprisingly, due to the combined effects of enhanced reverse transcription and defects in nuclear entry, two separate CA mutants that form hyperstable cores induced innate immune sensing more potently than destabilizing CA mutations. At low concentrations that allowed the accumulation of reverse transcription products, CA-targeting compounds GS-CA1 and lenacapavir measurably impacted CA lattice stability in cells and modestly enhanced innate immune sensing of HIV. Interestingly, innate immune activation observed with viruses containing unstable cores was abolished by low doses of lenacapavir. Innate immune activation observed with both hyperstable and unstable CA mutants was dependent on the cGAS-STING DNA-sensing pathway and reverse transcription. Overall, our findings demonstrate that CA lattice stability and reverse transcription are finely balanced to support reverse transcription and minimize cGAS-STING-mediated sensing of the resulting viral DNA. IMPORTANCE In HIV-1 particles, the dimeric RNA genome and associated viral proteins and enzymes are encased in a proteinaceous lattice composed of the viral capsid protein. Herein, we assessed how altering the stability of this capsid lattice through orthogonal genetic and chemical approaches impacts the induction of innate immune responses. Specifically, we found that decreasing capsid lattice stability results in more potent sensing of viral reverse transcription products, but not the genomic RNA, in a cGAS-STING-dependent manner. The recently developed capsid inhibitors lenacapavir and GS-CA1 enhanced the innate immune sensing of HIV-1. Unexpectedly, due to increased levels of reverse transcription and cytosolic accumulation of the resulting viral cDNA, capsid mutants with hyperstable cores also resulted in the potent induction of type I interferon-mediated innate immunity. Our findings suggest that HIV-1 capsid lattice stability and reverse transcription are finely balanced to minimize exposure of reverse transcription products in the cytosol of host cells.

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Emergence of Compensatory Mutations Reveals the Importance of Electrostatic Interactions between HIV-1 Integrase and Genomic RNA

Cited by 10 publications

References 105 publications

Oligomeric HIV-1 Integrase Structures Reveal Functional Plasticity for Intasome Assembly and RNA Binding

Oligomeric HIV-1 Integrase Structures Reveal Functional Plasticity for Intasome Assembly and RNA Binding

The C-terminal domain of RNase H and the C-terminus amino acid residue regulate virus release and autoprocessing of a defective HIV-1 possessing M50I and V151I changes in integrase

HIV-1 capsid stability and reverse transcription are finely balanced to minimize sensing of reverse transcription products via the cGAS-STING pathway

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