| The interaction of a T cell with a peptide-loaded antigen presenting cell (APC) induces the polarization of the cytoskeleton and signaling molecules towards the activating APC to a zone termed the immunological synapse (IS). One pathway implicated in cell polarity is the Rho family GTPase Cdc42 and its effector the Wiskott-Aldrich Syndrome protein (WASP). Both Cdc42 and WASP regulate cell polarity and actin dynamics in multiple cell types, suggesting that Cdc42 and WASP may be important regulators of IS formation in T cells.; In this work, we found that activated Cdc42 and WASP both localize to the IS upon T cell activation. Surprisingly, the interaction of WASP with Cdc42 is not responsible for WASP localization to the IS. Instead, interaction of the proline-rich region of WASP with the adaptor Nck is necessary and sufficient for WASP localization, showing that WASP activation can be separated from WASP recruitment. Lck and ZAP-70 phosphorylate SLP-76, which recruits both Vav and Nck. Vav is necessary for Cdc42 activation. At the IS, Vav activates Cdc42, leading to WASP activation, while Nck recruits WASP. This complex coordinates WASP recruitment and activation, leading to specific activation of WASP at the IS.; To study the effect of Cdc42 and WASP in T cell function, we found that mutants of Cdc42 block the ability of T cells to conjugate to APCs and polarize actin. Surprisingly, however, we found that although T cells from WASP-deficient mice show defects in IL2 production, WASP-deficient T cells conjugate normally to peptide-presenting APCs. Furthermore, WASP-deficient T cells polarize actin, MTOC, and the IS markers PKC&thetas; and talin normally. Although defects in the polarization of PKC&thetas; and talin were detected under low peptide stimulation conditions, the IL2 production defect in WASP-deficient T cells was seen at all peptide doses. These data strongly suggest that the role for WASP in IS formation is different from its role in IL2 production. Taken together, these results define the molecular pathway leading to WASP recruitment and activation at the IS, and show that WASP plays a role in IL2 production separate from actin regulation. |
