BCL-2 is phosphorylated and inactivated by an ASK1/Jun N-terminal protein kinase pathway normally activated at G(2)/M.
Multiple signal transduction pathways are capable of modifying BCL-2 family members to reset susceptibility to apoptosis. We used two-dimensional peptide mapping and sequencing to identify three residues (Ser70, Ser87, and Thr69) within the unstructured loop of BCL-2 that were phosphorylated in response to microtubule-damaging agents, which also arrest cells at G(2)/M. Changing these sites to alanine conferred more antiapoptotic activity on BCL-2 following physiologic death signals as well as paclitaxel, indicating that phosphorylation is inactivating. An examination of cycling cells enriched by elutriation for distinct phases of the cell cycle revealed that BCL-2 was phosphorylated at the G(2)/M phase of the cell cycle. G(2)/M-phase cells proved more susceptible to death signals, and phosphorylation of BCL-2 appeared to be responsible, as a Ser70Ala substitution restored resistance to apoptosis. We noted that ASK1 and JNK1 were normally activated at G(2)/M phase, and JNK was capable of phosphorylating BCL-2. Expression of a series of wild-type and dominant-negative kinases indicated an ASK1/Jun N-terminal protein kinase 1 (JNK1) pathway phosphorylated BCL-2 in vivo. Moreover, the combination of dominant negative ASK1, (dnASK1), dnMKK7, and dnJNK1 inhibited paclitaxel-induced BCL-2 phosphorylation. Thus, stress response kinases phosphorylate BCL-2 during cell cycle progression as a normal physiologic process to inactivate BCL-2 at G(2)/M.
Pubmed ID: 10567572 RIS Download
Amino Acid Sequence | Apoptosis | Binding Sites | CDC2 Protein Kinase | Cyclin B | Cyclin B1 | Enzyme Activation | G2 Phase | Humans | JNK Mitogen-Activated Protein Kinases | Jurkat Cells | MAP Kinase Kinase 7 | MAP Kinase Kinase Kinase 5 | MAP Kinase Kinase Kinases | MAP Kinase Signaling System | Microtubules | Mitogen-Activated Protein Kinase Kinases | Mitogen-Activated Protein Kinases | Mitosis | Molecular Sequence Data | Mutagenesis | Paclitaxel | Phosphorylation | Proto-Oncogene Proteins c-bcl-2 | Serine | Threonine