Dual regulation of T cell receptor-mediated signaling by oncogenic Cbl mutant 70Z

We previously showed that an oncogenic Cbl mutant (70Z) is constitutively a ctive in transcriptional activation of nuclear factor at activated T cells (NFAT), However, the mechanism underlying this effect remains unclear. Here we analyzed the effects of 70Z mutations at an amino-terminal loss of func tion site (Gly-306) and at carboxyl-terminal potential tyrosine or serine p hosphorylation sites on association with signaling proteins and on NFAT act ivation. Mutation at Gly-306 of 70Z disrupted its association with Zap-70 a nd almost completely abolished its ability to induce NFAT activation under basal and ionomycin-stimulated conditions. However, mutations at potential tyrosine or serine phosphorylation sites had little effect. In fact, expres sion of 70Z with Tyr-700, Tyr-731, or Tyr-774 mutated to Phe increased NFAT activity in comparison with unmutated 70Z. These findings suggest that an amino terminus-mediated interaction of 70Z with Zap-70 plays a positive rol e and that a carboxyl terminus-mediated, phosphotyrosine-dependent interact ion with their binding proteins plays a negative role in 70Z-mediated NFAT activation. In support of this notion are the observations that 70Z reduced T cell receptor-induced NFAT activation and that wild-type Cbl further inh ibited this event, suggesting that both 70Z and wild-type Cbl employ a simi lar mechanism by which Cbl proteins dually regulate T cell receptor-mediate d signaling.