THE ROLE OF P21(RAS) IN CD28 SIGNAL-TRANSDUCTION - TRIGGERING OF CD28WITH ANTIBODIES, BUT NOT THE LIGAND B7-1, ACTIVATES P21(RAS)

CD28 is a 44-kD homodimer expressed on the surface of the majority of human T cells that provides an important costimulus for T cell activat ion. The biochemical basis of the CD28 accessory signals is poorly und erstood. Triggering of the T cell antigen receptor (TCR) activates the p21(ras) proteins. Here we show that ligation of CD28 by a monoclonal antibody (mAb) also stimulates p21(ras) and induces Ras-dependent eve nts such as stimulation of the microtubule-associated protein (MAP) ki nase ERK2 and hyperphosphorylation of Raf-1. One physiological ligand for CD28 is the molecule B7-1. In contrast to the effect of CD28 mAb, the present studies show that interactions between CD28 and B7-1 do no t stimulate p21(ras) signaling pathways. Two substrates for TCR-regula ted protein tyrosine kinases (PTKs) have been implicated in p21(ras) a ctivation in T cells: p95(vav) and a 36-kD protein that associates wit h a complex of Grb2 and the Ras exchange protein Sos. Triggering CD28 with both antibodies and B7-1 activates cellular PTKs, and we have exp loited the differences between antibodies and B7-1 for p21(ras) activa tion in an attempt to identify critical PTK-controlled events for Ras activation in T cells. The data show that antibodies against TCR or CD 28 induce tyrosine phosporylation of both Vav and p36. B7-1 also induc es Vav tyrosine phosphorylation but has no apparent effect on tyrosine phosphorylation of the Grb2-associated p36 protein. The intensity of the Vav tyrosine phosphorylation is greater in B7-1 than in TCR-stimul ated cells. Moreover the kinetics of Vav tyrosine phosphorylation is p rolonged in the B7-1-stimulated cells. These studies show that for CD2 8 signaling, the activation of p21(ras) correlates more closely with p 36 tyrosine phosphorylation than with Vav tyrosine phosphorylation. Ho wever, the experiments demonstrate that Vav is a major substrate for B 7-activated PTKs and hence could be important in CD28 signal transduct ion pathway.