The Gene Encoding the Antisense Protein ASP of HIV-1: Origin, Distribution and Maintenance

Human Immunodeficiency Virus Type 1 (HIV-1), the causative agent of the acquired immune deficiency syndrome (AIDS), originated from zoonotic transmissions of simian immunodeficiency viruses (SIVs) infecting African great apes, following complex cross-species transmission events and virus-host co-evolution. These processes were accompanied by multiple viral adaptations, particularly within structural and accessory genes, enabling evasion of host restriction factors and long-term viral persistence. In 1988, an antisense open reading frame (ORF) overlapping the env gene was proposed and subsequently confirmed by the identification of antisense transcripts and the antisense protein (ASP). An "intact" ASP ORF (defined as >150 codons) is predominantly conserved in pandemic HIV-1 group M viruses and shows evidence of positive selection, suggesting a selective advantage. Increasing evidence supports the hypothesis that the asp gene emerged de novo during the evolution of group M and contributed to viral adaptation and global spread in humans. This review combines a narrative review of the literature with original in silico analyses of HIV-1 and SIV sequences retrieved from the Los Alamos National Laboratory database. We systematically reassessed the distribution, length variability and conservation of the ASP ORF across HIV-1 groups (M, N, O, P), subtypes, circulating recombinant forms (CRFs), unique recombinant forms (URFs) and related SIV lineages. Our updated analyses confirmed the strong association between the presence of an "intact" ASP ORF and pandemic HIV-1 group M lineages, while revealing rare but notable antisense ORFs in selected SIVcpz and SIVgor strains. By integrating evolutionary, epidemiological and sequence-based evidence, we aim to clarify the origin and maintenance of the ASP ORF and to contextualize its emergence within the broader framework of overlapping gene evolution, de novo gene birth and the selective pressures shaping viral fitness and pandemic potential.