Abstract

HIV-1 is a lentivirus that establishes a very intimate relationship with its host cell. At every step in its life cycle, this virus exploits host cell machinery for its own purposes. For example, during egress, the HIV-1 Gag protein recruits the cellular ESCRT (Endosomal Sorting Complex Required for Transport) machinery, in order to facilitate the final pinching off process. ESCRT proteins normally function in the cellular endocytic pathway, where they mediate the sorting of transmembrane proteins such as cell surface receptors into the lumen of the multvesicular body (MVB) prior to lysosomal degradation. The impact of viral exploitation of cellular ESCRT proteins on receptor downregulation is not known.

One example of a cell surface receptor that has previously been shown to require ESCRT machinery for lysosomal degradation is the Epidermal Growth Factor Receptor (EGFR). In this study, we addressed the effect of the expression of HIV-1 Gag proteins on lysosomal degradation of EGFR, using confocal microscopy. We found that ligand-induced EGFR downregulation was attenuated in HIV-1 Gag expressing cells. This observation indicates that HIV-1 Gag functionally depletes the cellular machinery required for EGFR downregulation.

In order to determine whether the endocytic trafficking of other cellular transmembrane proteins and receptors are similarly modulated by HIV-1 Gag, we studied the lysosomal downregulation of CD4 and CXCR4. CD4 and CXCR4 function as the receptor and co-receptor, respectively for the entry of HIV-1 into target cells. Regulation of cellular levels of these two proteins is critically important for HIV-1 pathogenesis. We first determined whether ESCRT proteins participate in the lysosomal downregulation of these proteins using siRNA, confocal microscopy and metabolic labeling techniques. We found that CXCR4 downregulation was ESCRT-I-dependent, while CD4 downregulation was not. HIV-1 Gag attenuated CXCR4 but not CD4 downregulation. Based on these observations, we conclude that HIVA Gag selectively modulates only ESCRT-dependent receptor-downregulation pathways in the cell.

In summary, this work establishes that HIV-1 Gag functionally depletes ESCRT complexes during egress, thereby attenuating ESCRT-dependent receptor-downregulation pathways and contributing to the altered physiology observed in HIVA infected cells.