No abstract

Growth factor deprivation is a physiological mechanism to regulate cell death. We utilize an interleukin‐2 (IL‐2)‐dependent murine T‐cell line to identify proteins that interact with Bad upon IL‐2 stimulation or deprivation. Using the yeast two‐hybrid system, glutathione S‐transferase (GST) fusion proteins and co‐immunoprecipitation techniques, we found that Bad interacts with protein phosphatase 1α (PP1α). Serine phosphorylation of Bad is induced by IL‐2 and its dephosphorylation correlates with appearance of apoptosis. IL‐2 deprivation induces Bad dephosphorylation, suggesting the involvement of a serine phosphatase. A serine/threonine phosphatase activity, sensitive to the phosphatase inhibitor okadaic acid, was detected in Bad immunoprecipitates from IL‐2‐stimulated cells, increasing after IL‐2 deprivation. This enzymatic activity also dephosphorylates in vivo 32P‐labeled Bad. Treatment of cells with okadaic acid blocks Bad dephosphorylation and prevents cell death. Finally, Ras activation controls the catalytic activity of PP1α. These results strongly suggest that Bad is an in vitro and in vivo substrate for PP1α phosphatase and that IL‐2 deprivation‐induced apoptosis may operate by regulating Bad phosphorylation through PP1α phosphatase, whose enzymatic activity is regulated by Ras.