Role for non-proteolytic control of M-phase-promoting factor activity at M-phase exit

• Published in: PloS one Vol. 2; no. 2; p. e247
• Main Authors: D'Angiolella, Vincenzo, Palazzo, Luca, Santarpia, Concetta, Costanzo, Vincenzo, Grieco, Domenico
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 28.02.2007; Public Library of Science (PLoS)
• Subjects: Anaphase; Anaphase-promoting complex; Animals; Biochemistry/Chemical Biology of the Cell; Biomedical materials; Biotechnology; Ca2+/calmodulin-dependent protein kinase II; Calcium; Calcium binding proteins; Calcium-binding protein; Calmodulin; CDC2 Protein Kinase - antagonists & inhibitors; CDC2 Protein Kinase - chemistry; CDC2 Protein Kinase - genetics; CDC2 Protein Kinase - physiology; cdc25 Phosphatases - physiology; Cell Biology/Cell Growth and Division; Cell cycle; Cell Cycle Proteins - physiology; Cyclin B; Cyclin B1; Cyclin B1 - chemistry; Cyclin B1 - genetics; Cyclin B1 - physiology; Deactivation; Deoxyribonucleic acid; Developmental Biology/Cell Differentiation; DNA; Eggs; Enzyme Activation; Female; Humans; Inactivation; Kinases; M-phase promoting factor; Meiosis; Metaphase - physiology; Mitosis; Molecular Biology/Chromatin Structure; Molecular Biology/Chromosome Structure; Molecular Biology/DNA; Molecular Biology/DNA Repair; Morphogenesis; Oncology; Oocytes - cytology; Phosphatases; Phosphorylation; Phosphothreonine - metabolism; Phosphotyrosine - metabolism; Physiological aspects; Protein Interaction Mapping; Protein kinase A; Protein kinases; Protein Processing, Post-Translational; Protein-Tyrosine Kinases - physiology; Proteolysis; Recombinant Fusion Proteins - physiology; Regulation; Surgical implants; Ubiquitin; Xenopus; Xenopus laevis; Xenopus Proteins - physiology; Italy
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0000247

M-phase Promoting Factor (MPF; the cyclin B-cdk 1 complex) is activated at M-phase onset by removal of inhibitory phosphorylation of cdk1 at thr-14 and tyr-15. At M-phase exit, MPF is destroyed by ubiquitin-dependent cyclin proteolysis. Thus, control of MPF activity via inhibitory phosphorylation is believed to be particularly crucial in regulating transition into, rather than out of, M-phase. Using the in vitro cell cycle system derived form Xenopus eggs, here we show, however, that inhibitory phosphorylation of cdk1 contributes to control MPF activity during M-phase exit. By sampling extracts at very short intervals during both meiotic and mitotic exit, we found that cyclin B1-associated cdk1 underwent transient inhibitory phosphorylation at tyr-15 and that cyclin B1-cdk1 activity fell more rapidly than the cyclin B1 content. Inhibitory phosphorylation of MPF correlated with phosphorylation changes of cdc25C, the MPF phosphatase, and physical interaction of cdk1 with wee1, the MPF kinase, during M-phase exit. MPF down-regulation required Ca(++)/calmodulin-dependent kinase II (CaMKII) and cAMP-dependent protein kinase (PKA) activities at meiosis and mitosis exit, respectively. Treatment of M-phase extracts with a mutant cyclin B1-cdk1AF complex, refractory to inhibition by phosphorylation, impaired binding of the Anaphase Promoting Complex/Cyclosome (APC/C) to its co-activator Cdc20 and altered M-phase exit. Thus, timely M-phase exit requires a tight coupling of proteolysis-dependent and proteolysis-independent mechanisms of MPF inactivation.