CONICET | Buscador de Institutos y Recursos Humanos

The acrosome reaction (AR) is a regulated calcium-dependent exocytosis necessary for fertilization. The HIV-1 transactivating protein (Tat) is released by infected cells and extracellular Tat enters uninfected cells by endocytosis inducing toxic effects. The aim of this work was to determine if HIV-1 Tat was able to enter a non-endocytic cell like the sperm thereby affecting gamete function. We incubated spermatozoa with recombinant wild type Tat (WT-Tat). WB and IFI assays demonstrated that, at physiological concentrations, Tat penetrates sperm membranes. To elucidate the mechanisms involved in Tat internalization we challenged sperm with Tat mutants. The W11 residue is required in this process. Exocytosis experiments demonstrated that WT-Tat inhibited progesterone (Pg)-induced AR. Tat binds phosphatidylinositol 4,5-bisphosphate (PIP2) with high affinity and our group has shown that PIP2 plays a key role in the sperm exocytic cascade. Additionally, we tested a Tat mutant unable to bind PIP2. This mutant did not affect sperm exocytosis. We rescued Tat-induced inhibition of secretion by adding PIP2. This suggests that Tat is sequestering PIP2. We assumed that Pg-induced AR inhibition will occur due to lack of IP3 synthesis. To test this argument, we resorted to the agonist of IP3 receptors, adenophostin that rescued Pg-induced exocytosis after Tat inhibition. This result was confirmed by calcium measurements. These findings suggest that Tat requires the W11 residue to penetrate the plasma membrane and once inside the sperm a strong interaction with PIP2 abolishes the AR. Our findings may contribute to elucidate unsolved issues concerning male subfertility in HIV patients.