This study demonstrates that human immunodeficiency virus type 1 (HIV‐1) Tat protein amplifies the activity of tumor necrosis factor (TNF), a cytokine that stimulates HIV‐1 replication through activation of NF‐kappa B. In HeLa cells stably transfected with the HIV‐1 tat gene (HeLa‐tat cells), expression of the Tat protein enhanced both TNF‐induced activation of NF‐kappa B and TNF‐mediated cytotoxicity. A similar potentiation of TNF effects was observed in Jurkat T cells and HeLa cells treated with soluble Tat protein. TNF‐mediated activation of NF‐kappa B and cytotoxicity involves the intracellular formation of reactive oxygen intermediates. Therefore, Tat‐mediated effects on the cellular redox state were analyzed. In both T cells and HeLa cells HIV‐1 Tat suppressed the expression of Mn‐dependent superoxide dismutase (Mn‐SOD), a mitochondrial enzyme that is part of the cellular defense system against oxidative stress. Thus, Mn‐SOD RNA protein levels and activity were markedly reduced in the presence of Tat. Decreased Mn‐SOD expression was associated with decreased levels of glutathione and a lower ratio of reduced:oxidized glutathione. A truncated Tat protein (Tat1‐72), known to transactivate the HIV‐1 long terminal repeat (LTR), no longer affected Mn‐SOD expression, the cellular redox state or TNF‐mediated cytotoxicity. Thus, our experiments demonstrate that the C‐terminal region of HIV‐1 Tat is required to suppress Mn‐SOD expression and to induce pro‐oxidative conditions reflected by a drop in reduced glutathione (GSH) and the GSH:oxidized GSH (GSSG) ratio.(ABSTRACT TRUNCATED AT 250 WORDS)

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Evidence exists that the human immunodeficiency virus-1 (HIV-1) transactivator Tat occurs extracellularly and is involved in the immunosuppression of non-HIV-1-infected T cells of acquired immunodeficiency syndrome (AIDS) patients. The mechanism of this immunosuppressive activity of Tat has been controversially discussed. Interestingly, Tat binds to the T cell activation marker CD26, which has been shown to play a key role in the regulation of growth of lymphocytes and to inhibit its dipeptidyl peptidase IV (DP IV) activity. Here we show that the N-terminal nonapeptide MDPVDPNIE of Tat is a competitive inhibitor of DP IV and suppresses DNA synthesis of tetanus toxoid-stimulated peripheral blood mononuclear cells. Amino acid exchanges at positions 5 and 6 strongly weaken these effects. 1H nuclear magnetic resonance and molecular dynamics simulations of Tat(1-9), I5-Tat(1-9), and L6-Tat(1-9) suggest a similar backbone conformation for Tat(1-9) and L6-Tat(1-9). The solution conformation of I5-Tat(1-9) considerably differs from the other two. However, Tat(1-9) fits into our previously proposed active site model of DP IV in contrast to I5-Tat(1-9) and L6-Tat(1-9). Conformational alterations with regard to the parent peptide and spatial hindrances between these both compounds and DP IV can explain the loss of inhibitory activity. Our data suggest that the N-terminal residues of HIV-1 Tat do interact directly with the active site of DP IV and that DP IV does mediate Tat's immunosuppressive effects.

Recent data in the literature suggest that the HIV-1 Tat(I-86) protein exhibits immunosuppressive effects. Moreover, Tat was found to interact with dipeptidyl peptidase IV (DP IV), which is identical to the T cell activation marker CD26. Here we show that the N-terminal amino acid sequence of Tat is essential for the inhibition of DP IV-catalyzed IL-2(1-12) degradation. Nterminal modification of Tat with rhodamine prevented inhibition of enzymatic activity of DP IV as well as suppression of DNA synthesis of mitogen-stimulated human T cells. Moreover, natural peptides containing the X-X-Pro N-terminal motif of Tat also inhibited DP IV activity. These data suggest the existence of endogenous immunomodulatory oligopeptides which influence immune cell proliferation and differentiation via DP IV a~ does HIV-1 Tat.