The biological significance of the PTAP motif duplication in HIV-1 subtype C p6 Gag

HIV-1 subtype C demonstrates an ability to duplicate longer stretches of amino acids in p6 Gag leading to the creation of an additional PTAP motif necessary for viral packaging. The biological significance of PTAP motif duplication for HIV-1 has not been experimentally validated. In a longitudinal study, of two different clinical cohorts of select seropositive drug-naive subjects of India, we found that 8 of 50 subjects contained a completePTAP duplication in p6 Gag. Conventional and Next-generation sequencing ofsixprimary viral quasispecies at multiple time-points demonstrated that in a mixed infectionthe viral strains containing PTAP duplication could establish an absolute dominationon the viral strains that lack such a duplication. The domination of the double-PTAP viral strains over the genetically similar single-PTAPviral clonewas confirmed in the viral proliferation assay and the pairwise competition assay. The Gag protein containing PTAP duplicationdemonstrated a significantly enhanced interactionwith Tsg101in the proximity ligation assay. Additionally, Tsg101 overexpression demonstrated a biphasic effect on the subtype C viral release, an enhancing effect at low concentrationand an inhibitory effect only at higher concentrations in contrast to the uniform inhibitory effect onsubtype B viral strains. Importantly, we show that double-PTAP viral strains gain replication advantage by a mechanism other than enhanced viral budding. In summary, it appears that Gag of subtype C, unlike that of subtype B, is endowed with a propensity to use a relatively higher concentration of Tsg101 to gain replication fitness. Our results have implication for pathogenesis of HIV-1 especially subtype C.