The Antisense Protein ASP of HIV-1 Enhances Viral Entry in CD4+ T Cells

The negative strand of the human immunodeficiency virus-1 (HIV-1) proviral genome contains an antisense open reading frame encoding a protein (ASP) with no known homologs. The presence of immune responses to ASP in people living with HIV-1 (PLWH) demonstrates its expression in vivo. Further, the predicted hydrophobicity of ASP is consistent with its association with the plasma membrane and viral envelope. Despite this body of evidence, the role of ASP in HIV-1 replication remains unknown. In this report, we investigated the hypothesis that the presence of ASP on the viral surface enhances HIV-1 entry into target cells. We generated an ASP-knockout replication-competent HIV-1 molecular clone in the NL4-3 background, which we used to perform cell-cell fusion, viral entry, and viral replication assays. Our results suggest that the presence of ASP on the plasma membrane of infected cells and the envelope of HIV-1 virions enhances viral transmission. Overall, our studies provide first evidence that ASP plays a role in the HIV-1 replication cycle. Further investigation into these observations may lead to the identification of new HIV-1 vulnerabilities that may be the target of novel interventions.