Pleiotropic effects of immobilized versus soluble recombinant HIV-1 Tat protein on CD3-mediated activation, induction of apoptosis, and HIV-1 long terminal repeat transactivation in purified CD4+ T lymphocytes.

摘要

CD3 mAb and HIV-1 Tat protein co-immobilized on plastic were able to induce a strong proliferation of resting human CD4 T cells, cultured in a serum-free chemically defined medium. Blocking studies performed with heparin or peptides containing the RGD sequence demonstrated that the heparin-binding basic domain of Tat plays a predominant role in CD4+ T cell activation. Moreover, the enhanced proliferative response of CD4+ T cells to immobilized Tat appeared to be mediated by alpha 5, beta 1, and alpha v subunits of surface integrin receptors. In contrast, soluble Tat showed a dose-dependent inhibitory activity on the proliferative response of resting CD4+ T cells stimulated by CD3 mAb co-immobilized with Tat or fibronectin, but not with CD28 mAb. In transient transfection assays performed with an HIV-1 long terminal repeat (LTR)-chloramphenicol acetyltransferase (CAT) plasmid CD3 mAb co-immobilized with Tat or fibronectin or CD28 mAb significantly stimulated CAT activity over the background. On the other hand, while immobilized Tat alone had no effects on LTR transactivation, soluble Tat was able to transactivate LTR-CAT in a dose-dependent manner. When CD4+ T cells activated by CD3 mAb co-immobilized with Tat were recovered, cultured for 7 days with 25 U/ml recombinant IL-2, and given an additional activation signal by recross-linking CD3 mAb, a marked increase of apoptosis was observed with respect to cells not subjected to CD3 mAb recross-linking. While co-immobilized Tat plus CD3 mAb did not show any significant effect on activation-induced cell death, high concentrations of soluble Tat synergized with immobilized CD3 mAb in the induction of apoptosis.