Dataset Information

A tyrosine-based motif in the HIV-1 envelope glycoprotein tail mediates cell-type- and Rab11-FIP1C-dependent incorporation into virions.

ABSTRACT: Lentiviruses such as HIV-1 encode envelope glycoproteins (Env) with long cytoplasmic tails (CTs) that include motifs mediating interactions with host-cell-trafficking factors. We demonstrated recently that Rab11-family interacting protein 1C (FIP1C) is required for CT-dependent incorporation of Env into HIV-1 particles. Here, we used viruses bearing targeted substitutions within CT to map the FIP1C-dependent incorporation of Env. We identified YW795 as a critical motif mediating cell-type-dependent Env incorporation. Disruption of YW795 reproduced the cell-type-dependent particle incorporation of Env that had previously been observed with large truncations of CT. A revertant virus bearing a single amino acid change near the C terminus of CT restored wild-type levels of Env incorporation, Gag-Env colocalization on the plasma membrane, and viral replication. These findings highlight the importance of YW795 in the cell-type-dependent incorporation of Env and support a model of HIV assembly in which FIP1C/RCP mediates Env trafficking to the particle assembly site.

SUBMITTER: Qi M

PROVIDER: S-EPMC4475960 | biostudies-other | 2015 Jun

REPOSITORIES: biostudies-other

ACCESS DATA

Similar Datasets

Project description:The HIV-1 envelope glycoprotein complex (Env) is incorporated into developing particles at the plasma membrane (PM). The cytoplasmic tail (CT) of Env is known to play an essential role in particle incorporation, while the exact mechanisms underlying this function of the CT remain uncertain. Upon reaching the PM, trafficking signals in the CT interact with host cell endocytic machinery, directing Env into endosomal compartments within the cell. Prior studies have suggested that Env must traffic through the endosomal recycling compartment (ERC) in order for Env to return to the plasma membrane (PM) site of particle assembly. Expression of a truncated form of the ERC-resident trafficking adaptor Rab11-Family Interacting Proteins C (FIP1C) resulted in CT-dependent sequestration of Env in the condensed ERC, preventing recycling of Env to the PM. In this work, the motifs within the CT responsible for ERC localization of Env were systematically mapped. A small deletion encompassing the N-terminal portion of LLP3 eliminated ERC localization. Site-directed mutagenesis identified two tryptophan-based motifs (WE 790-791 and WW 796-797 ) within the N-terminus of LLP3 that were essential for ERC localization of Env. Mutant viruses bearing substitutions in these motifs were deficient in Env incorporation, with a corresponding loss of particle infectivity and a significant defect in replication in a spreading infection assay. These results identify two tryptophan-based motifs at the N-terminal portion of LLP3 that mediate ERC localization and Env incorporation, providing additional supporting evidence for the importance of cellular recycling pathways in HIV-1 particle assembly.ImportanceThe HIV-1 envelope glycoprotein (Env) is an essential component of the virus, and has an exceedingly long cytoplasmic tail (CT). Previous studies have suggested that trafficking signals in the CT interact with host factors to regulate the incorporation of Env into particles. One particular area of interest is termed lentiviral lytic peptide 3 (LLP3), as small deletions in this region have been shown to disrupt Env incorporation. In this study, we identify a small region within LLP3 that regulates how Env associates with cellular recycling compartments. Mutants that reduced or eliminated Env from the recycling compartment also reduced Env incorporation into particles. These findings emphasize the importance of two tryptophan motifs in LLP3 to the incorporation of Env into particles, and provide additional support for the idea that the CT interacts with host recycling pathways to determine particle incorporation.

Project description:The matrix (MA) domain of HIV-1 Gag plays key roles in virus assembly by targeting the Gag precursor to the plasma membrane and directing the incorporation of the viral envelope (Env) glycoprotein into virions. The latter function appears to be in part dependent on trimerization of the MA domain of Gag during assembly, as disruption of the MA trimer interface impairs Env incorporation. Conversely, many MA mutations that impair Env incorporation can be rescued by compensatory mutations in the trimer interface. In this study, we sought to investigate further the biological significance of MA trimerization by isolating and characterizing compensatory mutations that rescue MA trimer interface mutants with severely impaired Env incorporation. By serially propagating MA trimerization-defective mutants in T cell lines, we identified a number of changes in MA, both within and distant from the trimer interface. The compensatory mutations located within or near the trimer interface restored Env incorporation and particle infectivity and permitted replication in culture. The structure of the MA lattice was interrogated by measuring the cleavage of the murine leukemia virus (MLV) transmembrane Env protein by the viral protease in MLV Env-pseudotyped HIV-1 particles bearing the MA mutations and by performing crystallographic studies of in vitro-assembled MA lattices. These results demonstrate that rescue is associated with structural alterations in MA organization and rescue of MA domain trimer formation. Our data highlight the significance of the trimer interface of the MA domain of Gag as a critical site of protein-protein interaction during HIV-1 assembly and establish the functional importance of trimeric MA for Env incorporation.IMPORTANCE The immature Gag lattice is a critical structural feature of assembling HIV-1 particles, which is primarily important for virion formation and release. While Gag forms a hexameric lattice, driven primarily by the capsid domain, the MA domain additionally trimerizes where three Gag hexamers meet. MA mutants that are defective for trimerization are deficient for Env incorporation and replication, suggesting a requirement for trimerization of the MA domain of Gag in Env incorporation. This study used a gain-of-function, forced viral evolution approach to rescue HIV-1 mutants that are defective for MA trimerization. Compensatory mutations that rescue virus replication do so by restoring Env incorporation and MA trimer formation. This study supports the importance of MA domain trimerization in HIV-1 replication and the potential of the trimer interface as a therapeutic target.

Dataset Information

A tyrosine-based motif in the HIV-1 envelope glycoprotein tail mediates cell-type- and Rab11-FIP1C-dependent incorporation into virions.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets