Localization and functions of native and eGFP-tagged capsid proteins in HIV-1 particles

• Type of Material: Article
• Authors: Ashwanth Francis, Emory University; Anna Cereseto, University of Trento; Parmit K Singh, Dana-Farber Cancer Institute; Jiong Shi, Vanderbilt University Medical Center; Eric Poeschla, University of Colorado Denver; Alan N Engelman, Dana-Farber Cancer Institute; Christopher Aiken, Vanderbilt University Medical Center; Gregory B Melikyan, Emory University
• Language: English
• Date: 2022-08-01
• Publisher: PLoS
• Publication Version: Final Published Version
• Copyright Statement: © 2022 Francis et al
• License: Creative Commons Attribution 4.0 International
• Final Published Version (URL): http://dx.doi.org/10.1371/journal.ppat.1010754
• Title of Journal or Parent Work: PLoS Pathogens
• Volume: 18
• Issue: 8
• Start Page: e1010754
• End Page: e1010754
• Grant/Funding Information: This work was supported by NIH grants R21 AI145541 (to A.C.F.), R01 AI129862 (to G.B.M.), U54 AI150472 (to G.B.M. and A.C.F.), P50 AI150481 (to A.N.E. and C.A.), and R01 AI052014 (to A.N.E.). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
• Keywords: Active Transport, Cell Nucleus; HIV Infections; Virus Replication; Virion; Capsid; Capsid Proteins; HIV-1; Virus Uncoating; Humans
• Research Categories: Health Sciences, Pathology; Health Sciences, Medicine and Surgery

Abstract

In infectious HIV-1 particles, the capsid protein (CA) forms a cone-shaped shell called the capsid, which encases the viral ribonucleoprotein complex (vRNP). Following cellular entry, the capsid is disassembled through a poorly understood process referred to as uncoating, which is required to release the reverse transcribed HIV-1 genome for integration into host chromatin. Whereas single virus imaging using indirect CA labeling techniques suggested uncoating to occur in the cytoplasm or at the nuclear pore, a recent study using eGFP-tagged CA reported uncoating in the nucleus. To delineate the HIV-1 uncoating site, we investigated the mechanism of eGFP-tagged CA incorporation into capsids and the utility of this fluorescent marker for visualizing HIV-1 uncoating. We find that virion incorporated eGFP-tagged CA is effectively excluded from the capsid shell, and that a subset of the tagged CA is vRNP associated. These results thus imply that eGFP-tagged CA is not a direct marker for capsid uncoating. We further show that native CA co-immunoprecipitates with vRNP components, providing a basis for retention of eGFP-tagged and untagged CA by sub-viral complexes in the nucleus. Moreover, we find that functional viral replication complexes become accessible to integrase-interacting host factors at the nuclear pore, leading to inhibition of infection and demonstrating capsid permeabilization prior to nuclear import. Finally, we find that HIV-1 cores containing a mixture of wild-type and mutant CA interact differently with cytoplasmic versus nuclear pools of the CA-binding host cofactor CPSF6. Our results suggest that capsid remodeling (including a loss of capsid integrity) is the predominant pathway for HIV-1 nuclear entry and provide new insights into the mechanism of CA retention in the nucleus via interaction with vRNP components.