Membrane localization and function of Vav3 in T cells depend on its association with the adapter SLP-76.
The Vav family of guanine exchange factors plays a critical role in lymphocyte proliferation, cytoskeletal reorganization, and gene transcription upon immunoreceptor engagement. Although the role of Vav1 in T cells is well documented, the role of Vav3 is less clear. We investigated the subcellular localization of Vav3 during T cell activation. We report here that phosphorylation of Vav3 on tyrosine residue Tyr(173) is not required for T cell receptor (TCR)-induced Vav3 membrane translocation or immunological synapse (IS) recruitment, but mutation of this residue enhanced TCR-induced nuclear factor of activated T cells (NFAT) activation. However, Vav3 mutants either containing an Src homology 2 (SH2)-disabled point mutation (R697L) or lacking its SH3-SH2-SH3 domains were unable to bind SLP-76 did not translocate to the membrane or to the IS and furthermore failed to activate NFAT. Importantly, the membrane translocation of Vav3 was abrogated in Lck, ZAP-70, LAT, and SLP-76-deficient T cells, where Vav3 binding to SLP-76 was disrupted. Finally, we confirmed and underlined the critical role of Vav3 in NFAT activation by knocking down Vav3 expression in Vav1-deficient T cells. Altogether, our data show that TCR-induced association of Vav3 with SLP-76 is required for its membrane/IS localization and function.
Charvet
Céline
C
Division of Cell Biology, La Jolla Institute for Allergy and Immunology, San Diego, California 92121, USA. charvet@liai.org
Canonigo
Ann Janette
AJ
Billadeau
Daniel D
DD
Altman
Amnon
A
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D019556:COS Cells; D018797:Cell Cycle Proteins / Q000096:biosynthesis / Q000737:chemistry*; D002462:Cell Membrane / Q000378:metabolism; D049109:Cell Proliferation; D004268:DNA-Binding Proteins / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D017930:Genes, Reporter; D020662:Guanine Nucleotide Exchange Factors; D006801:Humans; D047468:Immunoprecipitation; D019169:Jurkat Cells; D008214:Lymphocytes / Q000378:metabolism; D018613:Microscopy, Confocal; D009154:Mutation; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins / Q000737:chemistry; D010750:Phosphoproteins / Q000737:chemistry*; D010766:Phosphorylation; D010957:Plasmids / Q000378:metabolism; D017354:Point Mutation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis / Q000737:chemistry*; D051567:Proto-Oncogene Proteins c-vav; D015398:Signal Transduction; D013347:Subcellular Fractions; D013601:T-Lymphocytes / Q000378:metabolism*; D013997:Time Factors; D014157:Transcription Factors / Q000737:chemistry; D014162:Transfection; D014443:Tyrosine / Q000737:chemistry; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D020662:Guanine Nucleotide Exchange Factors; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins; D010750:Phosphoproteins; D011518:Proto-Oncogene Proteins; D051567:Proto-Oncogene Proteins c-vav; C092947:SLP-76 signal Transducing adaptor proteins; D014157:Transcription Factors; C496618:VAV1 protein, human; C496621:VAV3 protein, human; D014443:Tyrosine