Identification of a phospholipase C-gamma1 (PLC-gamma1) SH3 domain-binding site in SLP-76 required for T-cell receptor-mediated activation of PLC-gamma1 and NFAT.
SLP-76 is an adapter protein required for T-cell receptor (TCR) signaling. In particular, TCR-induced tyrosine phosphorylation and activation of phospholipase C-gamma1 (PLC-gamma1), and the resultant TCR-inducible gene expression, depend on SLP-76. Nonetheless, the mechanisms by which SLP-76 mediates PLC-gamma1 activation are not well understood. We now demonstrate that SLP-76 directly interacts with the Src homology 3 (SH3) domain of PLC-gamma1. Structure-function analysis of SLP-76 revealed that each of the previously defined protein-protein interaction domains can be individually deleted without completely disrupting SLP-76 function. Additional deletion mutations revealed a new, 67-amino-acid functional domain within the proline-rich region 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-gamma1 and is required for TCR-mediated activation of Erk, PLC-gamma1, and NFAT (nuclear factor of activated T cells). The adjacent Gads-binding domain of SLP-76, also within the proline-rich region, mediates inducible recruitment of SLP-76 to a PLC-gamma1-containing complex via the recruitment of both PLC-gamma1 and Gads to another cell-type-specific adapter, LAT. Thus, TCR-induced activation of PLC-gamma1 entails the binding of PLC-gamma1 to both LAT and SLP-76, a finding that may underlie the requirement for both LAT and SLP-76 to mediate the optimal activation of PLC-gamma1.
Yablonski
D
D
Department of Pharmacology, Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Bat Galim, Haifa 31096, Israel.
Kadlecek
T
T
Weiss
A
A
D048868:Adaptor Proteins, Signal Transducing; D001665:Binding Sites; D002458:Cell Fractionation; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004789:Enzyme Activation; D005786:Gene Expression Regulation; D017930:Genes, Reporter / Q000235:genetics; D006801:Humans; D015151:Immunoblotting; D007527:Isoenzymes / Q000378:metabolism*; D008954:Models, Biological; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D051966:Phospholipase C gamma; D010750:Phosphoproteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D014157:Transcription Factors / Q000378:metabolism*; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism*; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D004268:DNA-Binding Proteins; D007527:Isoenzymes; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D010750:Phosphoproteins; D011948:Receptors, Antigen, T-Cell; D011993:Recombinant Fusion Proteins; C092947:SLP-76 signal Transducing adaptor proteins; D014157:Transcription Factors; D010738:Type C Phospholipases; D051966:Phospholipase C gamma