Biology subjects

Waheed, A. A.

Publications and source records attributed to Waheed, A. A..

5 recordsLinked to original sources

bioRxiv2026

Inhibitors of HIV-1 maturation modulate conformational dynamics and heterogeneity of the immature Gag lattice

HIV-1 maturation requires the viral protease to cleave the Gag polyprotein into its constituent domains. Subsequent conformational and morphological changes result in a mature, infectious virion. Prior studies support a hypothesis in which Gag conformational dynamics and the heterogeneity of the immature Gag lattice enable proteolytic and morphological maturation. We developed a single-molecule Forster resonance energy transfer imaging approach to probe, in real time, the conformational dynamics of individual Gag molecules within immature HIV-1 virions. Our results capture Gag's conformational landscape and the spatial heterogeneity of the immature lattice. We evaluated inhibitors of maturation to identify immature lattice features that are modulated to disrupt maturation. Maturation inhibitors that act through diverse mechanisms, including lenacapavir and compounds that alter viral membrane composition, arrest Gag dynamics and reduce the spatial heterogeneity of the lattice. These observations support a model in which Gag dynamics and immature lattice organization regulate HIV-1 maturation.

biophysics↗

bioRxiv2026

A mechanism-aware stoichiometry platform resolves functional viral thresholds and induces antiviral hypersensitivity

Antiviral discovery is frequently hindered by a stoichiometric blind spot--a lack of resolution regarding the functional density of viral enzymes required for replication. Traditional screening pipelines rely on target-based or phenotypic assays that cannot distinguish between simple molecular binding and the crossing of a functional stoichiometric cliff. Here, we present a mechanism-aware platform that resolves these enzymatic requirements with single-virion precision. By integrating quantitative cryo-electron microscopy and genomic validation with Monte Carlo modeling, we map the stoichiometric landscapes of HIV-1 Protease (PR) and Reverse Transcriptase (RT). We uncover a striking disparity in enzymatic demand: a high-redundancy buffering capacity for PR ([~]40 monomers) contrasted with a high-threshold requirement for RT ([~]95 subunits). We demonstrate that by systematically de-buffering the virion, our platform induces a state of antiviral hypersensitivity, enabling the detection of therapeutic activity in novel and clinical inhibitors that remain invisible to traditional workflows. Furthermore, this multiplexed profiling enables de novo target identification, as inhibitors trigger failure exclusively on their respective stoichiometric arms. This platform provides a deterministic roadmap for de-risking drug discovery and identifying viral sub-stoichiometric vulnerabilities.

biophysics↗

bioRxiv2024

PSGL-1 excludes HIV Env from virion surface through spatial hindrance involving structural folding of the decameric repeats (DR)

P-selectin glycoprotein ligand-1 (PSGL-1), a mucin-like surface glycoprotein, is primarily expressed on lymphoid and myeloid cells. PSGL-1 has recently been identified as an HIV restriction factor, blocking HIV infectivity mainly through virion incorporation that sterically hinders virion attachment to target cells. PSGL-1 also inhibits HIV Env incorporation into virions. However, the molecular mechanisms of PSGL-1-mediated Env exclusion remained unclear. Here, we investigated the role of PSGL-1s extracellular (EC) and intracellular (IC) domains in Env exclusion. We demonstrate that both EC and IC are important for Env exclusion; when EC was deleted, PSGL-1 completely lost its ability to inhibit Env incorporation, whereas when IC was deleted, PSGL-1 partially lost this activity. In addition, when the decameric repeats (DR) were deleted from EC, PSGL-1 also lost its ability to inhibit Env incorporation. Sequential DR deletion mutagenesis further demonstrated that a minimum of 9 DRs is necessary for Env exclusion. Molecular modeling of the DR structure revealed that PSGL-1 mutants with 7 or fewer DRs pose as an extended "rod-like" structure, whereas those with 9 or more DRs collapse into a "coil-like" structure that spatially excludes Env. Our studies suggest a model in which Env exclusion involves Gag-mediated PSGL-1 targeting to the virion assembly site where DR-mediated spatial exclusion blocks Env incorporation.

microbiology↗

bioRxiv2021

Mechanism of Viral Glycoprotein Targeting by Membrane-associated-RING-CH Proteins

An emerging class of cellular inhibitory proteins has been identified that targets viral glycoproteins. These include the membrane-associated RING-CH (MARCH) family of E3 ubiquitin ligases that, among other functions, downregulate cell-surface proteins involved in adaptive immunity. The RING-CH domain of MARCH proteins is thought to function by catalyzing the ubiquitination of the cytoplasmic tails (CTs) of target proteins, leading to their degradation. MARCH proteins have recently been reported to target retroviral envelope glycoproteins (Env) and vesicular stomatitis virus G glycoprotein (VSV-G). However, the mechanism of antiviral activity remains poorly defined. Here we show that MARCH8 antagonizes the full-length forms of HIV-1 Env, VSV-G, Ebola virus glycoprotein (EboV-GP), and the spike (S) protein of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) thereby impairing the infectivity of virions pseudotyped with these viral glycoproteins. This MARCH8-mediated targeting of viral glycoproteins requires the E3 ubiquitin ligase activity of the RING-CH domain. We observe that MARCH8 protein antagonism of VSV-G is CT dependent. In contrast, MARCH8-mediated targeting of HIV-1 Env, EboV-GP and SARS-CoV-2 S protein by MARCH8 does not require the CT, suggesting a novel mechanism of MARCH-mediated antagonism of these viral glycoproteins. Confocal microscopy data demonstrate that MARCH8 traps the viral glycoproteins in an intracellular compartment. We observe that the endogenous expression of MARCH8 in several relevant human cell types is rapidly inducible by type I interferon. These results help to inform the mechanism by which MARCH proteins exert their antiviral activity and provide insights into the role of cellular inhibitory factors in antagonizing the biogenesis, trafficking, and virion incorporation of viral glycoproteins. ImportanceViral envelope glycoproteins are an important structural component on the surface of enveloped viruses that direct virus binding and entry and also serve as targets for the host adaptive immune response. In this study, we investigate the mechanism of action of the MARCH family of cellular proteins that disrupt the trafficking and virion incorporation of viral glycoproteins across several virus families. This research provides novel insights into how host cell factors antagonize viral replication, perhaps opening new avenues for therapeutic intervention in the replication of a diverse group of highly pathogenic enveloped viruses.

immunology↗

bioRxiv2020

The Vpu-interacting protein ATP6V0C regulates expression of tetherin and HIV-1 release

The HIV-1 accessory protein Vpu enhances virus release by down-regulating cell surface expression of the host restriction factor tetherin. To further understand the role of host proteins in Vpu function, we carried out yeast two-hybrid screening and identified the V0 subunit C of vacuolar ATPase (ATP6V0C) as a Vpu-binding protein. To examine the role of ATP6V0C in Vpu-mediated tetherin degradation and HIV-1 release, we knocked down ATP6V0C expression in HeLa cells and observed that ATP6V0C depletion impairs Vpu-mediated tetherin degradation, resulting in a defect in HIV-1 release. We also observed that overexpression of ATP6V0C stabilizes tetherin expression. This stabilization is specific to ATP6V0C, as overexpression of another subunit of the vacuolar ATPase, ATP6V0C", had no effect on tetherin expression. ATP6V0C overexpression did not stabilize CD4, another target of Vpu-mediated degradation. Immunofluorescence localization studies showed that the ATP6V0C-stabilized tetherin is sequestered in a CD63- and LAMP1-positive intracellular compartment. These data demonstrate that the Vpu-interacting protein ATP6V0C plays a role in regulating tetherin expression and HIV-1 assembly and release.

microbiology↗