Figure 6.

Effect of Vav1 deficiency on PLCγ1 recruitment and activation. (A) Jurkat E6 cells (two left lanes) and Jurkat J.vav cells (two right lanes) were either stimulated with OKT3 for 1 min or left unstimulated and lysed. The lysates were resolved on SDS–PAGE and probed with either anti-phospho-PLCγ1-pY783 (upper blot) or anti-PLCγ1 (lower blot). The data are representative of two independent experiments. (B) Time course of the response in cytosolic calcium concentration obtained from the indicated cell lines after stimulation with OKT3. AU, arbitrary units. (C) Jurkat E6 or J.vav cells (1 × 10⁷) expressing PLCwt-YFP were either stimulated for 1 min with C305 or left unstimulated and lysed. PLCwt-YFP was precipitated with anti-GFP. The precipitates were resolved on SDS–PAGE and probed with anti-phosphotyrosine 4G10 antibody. The data are representative at least two independent experiments. (D) Jurkat J.vav cells expressing an indicated PLCγ1-YFP conjugate were seeded on stimulatory coverslips. Images obtained 1–1.5 min into the spreading process are shown. The size bar corresponds to 5 μm.