Mitosis persists in the absence of Cdk1 activity when proteolysis or protein phosphatase activity is suppressed.
Cellular transition to anaphase and mitotic exit has been linked to the loss of cyclin-dependent kinase 1 (Cdk1) kinase activity as a result of anaphase-promoting complex/cyclosome (APC/C)-dependent specific degradation of its cyclin B1 subunit. Cdk1 inhibition by roscovitine is known to induce premature mitotic exit, whereas inhibition of the APC/C-dependent degradation of cyclin B1 by MG132 induces mitotic arrest. In this study, we find that combining both drugs causes prolonged mitotic arrest in the absence of Cdk1 activity. Different Cdk1 and proteasome inhibitors produce similar results, indicating that the effect is not drug specific. We verify mitotic status by the retention of mitosis-specific markers and Cdk1 phosphorylation substrates, although cells can undergo late mitotic furrowing while still in mitosis. Overall, we conclude that continuous Cdk1 activity is not essential to maintain the mitotic state and that phosphatase activity directed at Cdk1 substrates is largely quiescent during mitosis. Furthermore, the degradation of a protein other than cyclin B1 is essential to activate a phosphatase that, in turn, enables mitotic exit.
Skoufias
Dimitrios A
DA
Institut de Biologie Structurale Jean-Pierre Ebel, Atomic Energy Commission/Centre National de la Recherche Scientifique, 38027 Grenoble, Cedex 1, France. dimitrios.skoufias@ibs.fr
Indorato
Rose-Laure
RL
Lacroix
Françoise
F
Panopoulos
Andreas
A
Margolis
Robert L
RL
D015075:2-Aminopurine / Q000031:analogs & derivatives / Q000494:pharmacology; D016203:CDC2 Protein Kinase / Q000037:antagonists & inhibitors*; D004396:Coloring Agents; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004347:Drug Interactions; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016650:Fluorescein-5-isothiocyanate; D019084:Fluorescent Antibody Technique, Indirect; D005456:Fluorescent Dyes; D045325:HCT116 Cells; D006367:HeLa Cells; D006801:Humans; D006868:Hydrolysis; D007769:Lactams / Q000494:pharmacology; D007976:Leupeptins / Q000494:pharmacology; D008938:Mitosis / Q000187:drug effects*; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors*; D011419:Propidium; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011687:Purines / Q000494:pharmacology; D000077546:Roscovitine
C471843:6-((3-chloro)anilino)-2-(isopropyl-2-hydroxyethylamino)-9-isopropylpurine; C477301:AM 114; D004396:Coloring Agents; D015853:Cysteine Proteinase Inhibitors; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D007769:Lactams; D007976:Leupeptins; C469937:N(2)-(2-aminocyclohexyl)-N(6)-(3-chlorophenyl)-9-ethyl-9H-purine-2,6-diamine; D047428:Protein Kinase Inhibitors; D011687:Purines; D000077546:Roscovitine; D011419:Propidium; D015075:2-Aminopurine; D016203:CDC2 Protein Kinase; D010749:Phosphoprotein Phosphatases; D016650:Fluorescein-5-isothiocyanate; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde