TRAF3 in T cells restrains negative regulators of LAT to promote TCR/CD28 signaling

Abstract The adaptor protein TRAF3 plays an important role in T cell biology. T cell conditional TRAF3−/− mice, despite normal T cell numbers, have poor responses to immunization and infection, and T cells from these mice have defective TCR/CD28 signaling. TRAF3 associates with, and enhances signaling by, the TCR/CD28 complex upon engagement of both TCR and CD28. The molecular mechanism by which TRAF3 interacts with and impacts TCR/CD28-mediated complexes and function remains an important knowledge gap. We investigated how TRAF3 is recruited to, and regulates, the TCR/CD28 complex. Our work revealed that TRAF3 recruitment did not require direct binding of TRAF3 to CD28; rather, TRAF3 associates with the CD28-binding protein Linker of activated T cells (LAT). The TRAF3-LAT association required the TRAF3 TRAF-C domain and a newly identified TRAF2/3 binding motif in LAT. TRAF3 enhanced TCR/CD28 signaling via inhibition of the LAT-associated negative regulatory protein Dok1, which is phosphorylated and inactivated by the tyrosine kinase Breast tumor kinase (Brk/PTK6). In TRAF3−/− T cells, there was increased plasma membrane localization of Protein tyrosine phosphatase 1B (PTP1B), which dephosphorylates Brk at an activating tyrosine residue, resulting in altered Brk activation. Consistent with these findings, inhibiting Brk activity in WT T cells inhibited basal and TCR/CD28-mediated Src family kinase activation, while inhibiting PTP1B activity in TRAF3−/− T cells rescued such activation. These results reveal a new mechanism for the enhancement of TCR/CD28-mediated signaling by TRAF3. We are currently investigating the role of TRAF3 in T cell metabolic reprogramming, which requires effective TCR/CD28 signaling.