FIGURE 1.

Stimulation of CTL clones with agents that increase intracellular Ca²⁺ concentration induce Pyk2 phosphorylation. A, AB.1 CTL clones were stimulated with the indicated concentration of ionomycin or DMSO as a carrier control for 10 min. The cells were lysed, and Pyk2 was immunoprecipitated from post-nuclear lysates. The immunoprecipitates were subjected to SDS-PAGE followed by immunoblottting with anti-phosphotyrosine. The membrane was then stripped and reprobed for Pyk2. B, samples of the post-nuclear lysates from A were immunoblotted for anti-phosphotyrosine. C, AB.1 cells were stimulated with ionomycin in the presence or absence of 4 mm EGTA. Pyk2 immunoprecipitates were probed with anti-phosphotyrosine then anti-Pyk2. D, AB.1 were treated with the indicated concentration of thapsigargin or DMSO as a carrier control for 10 min. Pyk2 immunoprecipitates were probed for phosphotyrosine and then reprobed for Pyk2. E, AB.1 cells were stimulated with 0.5 μm thapsigargin for 10 min in the presence or absence of 4 mm EGTA and Pyk2 immunoprecipitates were subjected to immunoblotting with anti-phosphotyrosine followed by anti-Pyk2.