Biology subjects

Hudait, A.

Publications and source records attributed to Hudait, A..

6 recordsLinked to original sources

bioRxiv2026

Mechanistic Insights into HIV-1 Capsid Interactions with CPSF6

A crucial stage of the HIV-1 life cycle is the docking of the viral capsid at, and its translocation through, the nuclear pore complex (NPC). During this process, the capsid interacts with a series of cellular host factors that regulate efficient nuclear entry. One such host factor is cleavage and polyadenylation specificity factor 6 (CPSF6), which plays a critical role in efficient HIV-1 nuclear entry and integration. Experimental evidence suggests that nucleoporins (NUPs), such as NUP153, initially engage the capsid, followed by CPSF6 binding and oligomerization during subsequent stages of nuclear import. However, the mechanistic basis by which CPSF6 binds to the capsid and subsequently oligomerizes remains poorly understood. To address this question, we have developed bottom-up coarse-grained (CG) models that explicitly simulate the process of CPSF6 binding and oligomerization on the HIV-1 capsid. Our simulations show that CPSF6 assembles into a mesh-like coating surrounding the capsid, consistent with experimental observations. Furthermore, we then demonstrate that CPSF6 binding is dependent on the intrinsic curvature of the capsid lattice. Finally, we investigate how perturbing CPSF6-CPSF6 interactions alters binding, oligomerization dynamics, and curvature dependence. Together, these findings provide new mechanistic insights into how CPSF6 regulates HIV-1 nuclear entry.

biophysics↗

bioRxiv2025

Lenacapavir-induced Lattice Hyperstabilization is Central to HIV-1 Capsid Failure at the Nuclear Pore Complex and in the Cytoplasm

Lenacapavir (LEN) is the first HIV-1 capsid inhibitor approved for clinical use. It inhibits multiple steps of the viral life cycle; however, the molecular details of the effect of LEN on capsid structure and the mechanistic steps of the inhibition are not understood. Recent studies show that intact cone-shaped capsids and capsids with LEN-induced breaks can dock at nuclear pore complexes (NPC), but only intact capsids enter the nucleus. In this work, we combined large-scale coarse-grained molecular dynamics simulations and live-cell imaging to investigate the stepwise mechanism of docking of LEN-treated capsids into the NPC. Capsids bound to substoichiometric concentrations of LEN can reach the NPC central channel. As the capsid advances to the nuclear end, lattice defects are formed at the pentamer-hexamer interface - primarily at the narrower end - leading to pentamer dissociation. Dissociation of pentamers is detrimental to capsid integrity, leading to both rupture of the narrow end and destabilization of the hexamer-hexamer interface. Structural analysis of LEN-capsid complexes in our simulations demonstrates heterogeneous hyperstabilization and loss of the essential pliability of the capsid protein lattice. Live-cell imaging of HIV-1 cores labeled with two different fluorescent markers showed that LEN-treated ruptured capsids were docked at the NPC but were not imported into the nucleus. We conclude that LEN contributes to the loss of capsid elasticity and integrity, inhibiting HIV-1 nuclear entry and replication. Our findings demonstrate that altering viral material properties can be an effective strategy for designing antiviral drugs that disrupt viral core nuclear entry.

biophysics↗

bioRxiv2024

Kinetic Implications of IP6 Anion Binding on the Molecular Switch of the HIV-1 Capsid Assembly

HIV-1 capsid proteins (CA) self-assemble into a fullerene-shaped capsid, enabling cellular transport and nuclear entry of the viral genome. A structural switch comprising the Thr-Val-Gly-Gly (TVGG) motif either assumes a disordered coil or a 310 helix conformation to regulate hexamer or pentamer assembly, respectively. The cellular polyanion inositol hexakisphosphate (IP6) binds to a positively charged pore of CA capsomers rich in arginine and lysine residues mediated by electrostatic interactions. Both IP6 binding and TVGG coil-to-helix transition are essential for pentamer formation. However, the connection between IP6 binding and TVGG conformational switch remains unclear. Using extensive atomistic simulations, we show that IP6 imparts structural order at the central ring, which results in multiple kinetically controlled events leading to the coil- to-helix conformational change of the TVGG motif. IP6 facilitates the helix-to-coil transition by allowing the formation of intermediate conformations. Our results identify the key kinetic role of IP6 in HIV-1 pentamer formation.

biophysics↗

bioRxiv2023

HIV-1 Capsid Shape, Orientation, and Entropic Elasticity Regulate Translocationinto the Nuclear Pore Complex

Nuclear import of the viral capsid is a critical step in the HIV-1 life cycle that serve to transport and release genomic material into the nucleus. Nuclear Pore Complex (NPC) allows passage of intact capsid, though mechanistic details of the process remain to be fully understood. Here we investigate the factors regulating HIV-1 capsid translocation into the NPC central channel using coarse-grained molecular dynamics simulations. We find that successful translocation is contingent on the compatibility of the capsid morphology and channel dimension and the proper orientation of the capsid approach to the channel. The central channel dynamically expands to allow capsid passage, demonstrating the pleomorphic nature of the channel necessary for transporting large cargoes. Structural analysis shows that stress induced by the central channel confinement and uncondensed internal genomic material generates correlated striated patterns of lattice disorder across the viral capsid surface which is an indicator of its lattice "elasticity". Our results suggest that the "elasticity" can aid the capsid to adapt to the stress and remain structurally intact during translocation. TeaserComputer Simulations identify capsid elasticity as a key factor for successful nuclear entry.

biophysics↗

bioRxiv2022

Organization of Upstream ESCRT Machinery at the HIV-1 Budding Site

In the late stages of the HIV-1 life cycle, membrane localization and self-assembly of the Gag polyproteins induce membrane deformation and budding. However, release of the immature virion requires direct interaction between Gag lattice and upstream ESCRT machinery at the budding site, followed by assembly of the downstream ESCRT-III factors, culminating in membrane scission. In this work, using "bottom-up" coarse-grained (CG) molecular dynamics (MD) simulations we investigated the interactions between Gag and different upstream ESCRT components to delineate the molecular organization of proteins at the membrane neck of the HIV-1 budding site. We developed CG models of upstream ESCRT proteins and HIV-1 structural protein Gag based on experimental structural data and extensive all-atom MD simulations. We find that ESCRT-I proteins bound to the immature Gag lattice can recruit multiple copies of ESCRT-II coating the membrane neck. ESCRT-I can effectively oligomerize to higher-order complexes both in absence of ESCRT-II and when multiple copies of ESCRT-II are localized at the bud neck. The ESCRT-I/II supercomplexes observed in our simulations exhibit predominantly extended conformations. Importantly, the ESCRT-I/II supercomplex modulates the membrane mechanical properties at the budding site by decreasing the overall Gaussian curvature of membrane neck. Our findings serve to elucidate a network of interactions between the upstream ESCRT machinery, immature Gag lattice, and membrane bud neck that regulate the protein assemblies and enable bud neck constriction.

biophysics↗

bioRxiv2020

A helical assembly of human ESCRT-I scaffolds reverse-topology membrane scission

The ESCRT complexes drive membrane scission in HIV-1 release, autophagosome closure, MVB biogenesis, cytokinesis, and other cell processes. ESCRT-I is the most upstream complex and bridges the system to HIV-1 Gag in virus release. The crystal structure of the headpiece of human ESCRT-I comprising TSG101:VPS28:VPS37B:MVB12A was determined, revealing an ESCRT-I helical assembly with a 12 molecule repeat. Electron microscopy confirmed that ESCRT-I subcomplexes form helical filaments in solution. Mutation of VPS28 helical interface residues blocks filament formation in vitro and autophagosome closure and HIV-1 release in human cells. Coarse grained simulations of ESCRT assembly at HIV-1 budding sites suggest that formation of a 12-membered ring of ESCRT-I molecules is a geometry-dependent checkpoint during late stages of Gag assembly and HIV-1 budding, and templates ESCRT-III assembly for membrane scission. These data show that ESCRT-I is not merely a bridging adaptor, but has an essential scaffolding and mechanical role in its own right.

biophysics↗