Functional cooperation between c-Cbl and Src-like adaptor protein 2 in the negative regulation of T-cell receptor signaling.
Adaptor proteins assemble multiprotein signaling complexes, enabling the transduction of intracellular signals. While many adaptor proteins positively regulate signaling in this manner, a subgroup of adaptors function as negative regulators. Here we report the identification of a hematopoiesis-specific adaptor protein that we have designated Src-like adaptor protein 2 (SLAP-2). SLAP-2 is most closely related to SLAP and contains a Src homology 3 (SH3) domain and an SH2 domain, as well as an amino-terminal myristoylation site that mediates SLAP-2 association with membranes. Following stimulation of primary thymocytes with anti-CD3 and anti-CD28, SLAP-2 coimmunoprecipitates with tyrosine-phosphorylated c-Cbl and an unidentified protein of approximately 72 kDa. In activated Jurkat T cells, SLAP-2 also binds an additional 70-kDa phosphoprotein, identified as ZAP-70. Binding of SLAP-2 to both p72 and ZAP-70 is dependent on its SH2 domain, while c-Cbl interacts with the carboxy-terminal region. Overexpression of wild-type SLAP-2 alone or in combination with c-Cbl in Jurkat T cells leads to inhibition of T-cell antigen receptor-induced activation of nuclear factor of activated T cells. The inhibitory effect of SLAP-2 requires the carboxy-terminal c-Cbl binding region. Expression of SLAP-2 with SYK or ZAP-70 in COS cells or Jurkat T cells causes the degradation of these kinases, and SLAP-2 overexpression in Jurkat T cells reduces the surface expression of CD3. These results suggest that the mechanism of action of SLAP-2 and the related protein SLAP is to promote c-Cbl-dependent degradation of the tyrosine kinases SYK and ZAP-70 and down-regulation of CD3 at the cell surface.
Loreto
Michael P
MP
Department of Medical Biophysics, University of Toronto and Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada M5G 1X8.
Berry
Donna M
DM
McGlade
C Jane
CJ
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D018106:CD28 Antigens / Q000276:immunology; D017252:CD3 Complex / Q000378:metabolism; D002460:Cell Line; D003001:Cloning, Molecular; D017124:Conserved Sequence; D004268:DNA-Binding Proteins / Q000378:metabolism; D015536:Down-Regulation; D004792:Enzyme Precursors / Q000378:metabolism; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D019169:Jurkat Cells; D051379:Mice; D008969:Molecular Sequence Data; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D016315:Proto-Oncogene Proteins c-abl / Q000235:genetics / Q000378:metabolism*; D016392:Proto-Oncogene Proteins pp60(c-src) / Q000235:genetics* / Q000378:metabolism*; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism*; D015398:Signal Transduction; D000072377:Syk Kinase; D013601:T-Lymphocytes / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism; D051746:ZAP-70 Protein-Tyrosine Kinase
D048868:Adaptor Proteins, Signal Transducing; D018106:CD28 Antigens; D017252:CD3 Complex; C121704:CD3 antigen, zeta chain; D004268:DNA-Binding Proteins; D004792:Enzyme Precursors; D047908:Intracellular Signaling Peptides and Proteins; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D011948:Receptors, Antigen, T-Cell; C441328:SLA2 protein, human; C486695:Sla2 protein, mouse; D014157:Transcription Factors; D011505:Protein-Tyrosine Kinases; D016315:Proto-Oncogene Proteins c-abl; D016392:Proto-Oncogene Proteins pp60(c-src); C000606896:SYK protein, human; D000072377:Syk Kinase; C000606897:Syk protein, mouse; D051746:ZAP-70 Protein-Tyrosine Kinase; C497261:ZAP70 protein, human; C497262:Zap70 protein, mouse