Identification of a phospholipase C-gamma 1 (PLC-gamma 1) SH3 domain-binding site in SLP-76 required for T-cell receptor-mediated activation of PLC-gamma 1 and NFAT

SLP-76 is an adapter protein required for T-cell receptor (TCR) signaling. In particular, TCR-induced tyrosine phosphorylation and activation of phosp holipase C-gammal (PLC-gammal), and the resultant TCR-inducible gene expres sion, depend on SLP-76. Nonetheless, the mechanisms by which SLP-76 mediate s PLC-gamma1 activation are not well understood. We now demonstrate that SL P-76 directly interacts with the Src homology 3 (SH3) domain of PLC-gammal. Structure-function analysis of SLP-76 revealed that each of the previously defined protein-protein interaction domains can be individually deleted wi thout completely disrupting SLP-76 function. Additional deletion mutations revealed a new, 67-amino-acid functional domain within the proline-rich reg ion of SLP-76, which we have termed the P-1 domain. The P-1 domain mediates a constitutive interaction of SLP-76 with the SH3 domain of PLC-gammal and is required for TCR-mediated activation of Erk, PLC-gammal, and NFAT (nucl ear factor of activated T cells). The adjacent Gads-binding domain of SLP-7 6, also within the proline-rich region, mediates inducible recruitment of S LP-76 to a PLC-gammal-containing complex via the recruitment of both PLC-ga mmal and Gads to another cell-type-specific adapter, LAT. Thus, TCR-induced activation of PLC-gammal entails the binding of PLC-gammal to both LAT and SLP-76, a finding that may underlie the requirement for both LAT and SLP-7 6 to mediate the optimal activation of PLC-gammal.