SH2 domain containing leukocyte phosphoprotein of 76-kDa (SLP-76) feedback regulation of ZAP-70 microclustering.
T cell receptor (TCR) signaling involves CD4/CD8-p56lck recruitment of ZAP-70 to the TCR receptor, ZAP-70 phosphorylation of LAT that is followed by LAT recruitment of the GADS-SLP-76 complex. Back regulation of ZAP-70 by SLP-76 has not been documented. In this paper, we show that anti-CD3 induced ZAP-70 cluster formation is significantly reduced in the absence of SLP-76 (i.e., J14 cells) and in the presence of a mutant of SLP-76 (4KE) in Jurkat and primary T cells. Both the number of cells with clusters and the number of clusters per cell were reduced. This effect was not mediated by SLP-76 SH2 domain binding to ZAP-70 because SLP-76 failed to precipitate ZAP-70 and an inactivating SH2 domain mutation (i.e., R448L) on SLP-76 4KE did not reverse the inhibition of ZAP-70 clustering. Mutation of R448 on WT SLP-76 still supported ZAP-70 clustering. Intriguingly, by contrast, LAT clustering occurred normally in the absence of SLP-76, or the presence of 4KE SLP-76 indicating that this transmembrane adaptor can operate independently of ZAP-70-GADS-SLP-76. Our findings reconfigure the TCR signaling pathway by showing SLP-76 back-regulation of ZAP-70, an event that could ensure that signaling components are in balance for optimal T cell activation.
Cell Signalling Section, Department of Pathology, University of Cambridge, Cambridge CB2 1QP, United Kingdom.
Hebin
H
Liu
Marco A
MA
Purbhoo
Daniel M
DM
Davis
Christopher E
CE
Rudd
D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D019943:Amino Acid Substitution; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D025461:Feedback, Physiological; D006801:Humans; D019169:Jurkat Cells; D008213:Lymphocyte Activation; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism; D046912:Multiprotein Complexes; D016297:Mutagenesis, Site-Directed; D010750:Phosphoproteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D010766:Phosphorylation; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D015398:Signal Transduction; D013601:T-Lymphocytes / Q000276:immunology* / Q000378:metabolism*; D051746:ZAP-70 Protein-Tyrosine Kinase / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism*; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D017931:DNA Primers; C409587:GRAP2 protein, human; C110810:LAT protein, human; D008565:Membrane Proteins; D046912:Multiprotein Complexes; D010750:Phosphoproteins; D011948:Receptors, Antigen, T-Cell; D011994:Recombinant Proteins; C092947:SLP-76 signal Transducing adaptor proteins; D051746:ZAP-70 Protein-Tyrosine Kinase; C497261:ZAP70 protein, human