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<%AbstractTime%> S4.
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TCR-Independent Activation of T-Cell Signaling by HIV-1 Nef
H. Renkema, A. Manninen, and K. Saksela*
Inst. of Med. Technology, Univ. of Tampere, Finland
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Background: An important function of the HIV/SIV pathogenicity factor Nef is to promote host cell activation. A role of Nef in triggering or enhancing signal transduction via the T-cell receptor (TCR) has recently received significant attention. Our recent studies, however, have revealed molecular mechanisms by which Nef can activate T-cell signal transduction independently of TCR.
Methods: A panel of molecular and cell biology techniques has been used to study function of Nef in cell culture.
Results: We found that calcium/calcineurin-dependent activation of nuclear factor of activated T cells (NFAT) is initiated downstream of TCR, and requires the function of the endoplasmic reticulum-resident IP3 receptor (IP3R), but is not mediated via increased phospholipase-C?1-catalyzed production of IP3, nor via depletion of IP3-regulated intracellular calcium stores. Nef could be co-precipitated with endogenous IP3R from Nef-transfected Jurkat T cells as well as from HIV-infected primary human peripheral mononuclear cell cultures. Thus, we conclude that TCR-independent activation of T-cell calcium signaling is mediated via binding of Nef to IP3R, which leads to aberrant opening of calcium influx channels independently of the filling state of intracellular calcium stores. Another cellular target utilized by Nef to activate host cells that we have identified is the p21-activated kinase-2 (PAK2). Our recent work has focused on molecular characterization of the Nef/PAK2 interaction. These data indicate that the ability of Nef to activate and associate with PAK2 are 2 separate functions of Nef. The positive effect of Nef on PAK2 activation is indirect, and unlike the natural PAK activators, GTP-bound forms of Cdc42 and Rac, Nef can only bind to PAK2 molecules that have already undergone conformational activation. Consequently, Nef-associated PAK2 differs from the majority of PAK2 in cells by having a much higher intrinsic catalytic activity, an exclusive membrane localization, and by being a target for tyrosine phosphorylation.
Conclusions: Nef has several strategies to promote T-cell activation independently of TCR, which might be particularly important at later stages of infection of a T lymphocyte when TCR function and cell surface expression have been down-modulated.
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