A conserved cyclin-binding domain determines functional interplay between anaphase-promoting complex-Cdh1 and cyclin A-Cdk2 during cell cycle progression.
Periodic activity of the anaphase-promoting complex (APC) ubiquitin ligase determines progression through multiple cell cycle transitions by targeting cell cycle regulators for destruction. At the G(1)/S transition, phosphorylation-dependent dissociation of the Cdh1-activating subunit inhibits the APC, allowing stabilization of proteins required for subsequent cell cycle progression. Cyclin-dependent kinases (CDKs) that initiate and maintain Cdh1 phosphorylation have been identified. However, the issue of which cyclin-CDK complexes are involved has been a matter of debate, and the mechanism of how cyclin-CDKs interact with APC subunits remains unresolved. Here we substantiate the evidence that mammalian cyclin A-Cdk2 prevents unscheduled APC reactivation during S phase by demonstrating its periodic interaction with Cdh1 at the level of endogenous proteins. Moreover, we identified a conserved cyclin-binding motif within the Cdh1 WD-40 domain and show that its disruption abolished the Cdh1-cyclin A-Cdk2 interaction, eliminated Cdh1-associated histone H1 kinase activity, and impaired Cdh1 phosphorylation by cyclin A-Cdk2 in vitro and in vivo. Overexpression of cyclin binding-deficient Cdh1 stabilized the APC-Cdh1 interaction and induced prolonged cell cycle arrest at the G(1)/S transition. Conversely, cyclin binding-deficient Cdh1 lost its capability to support APC-dependent proteolysis of cyclin A but not that of other APC substrates such as cyclin B and securin Pds1. Collectively, these data provide a mechanistic explanation for the mutual functional interplay between cyclin A-Cdk2 and APC-Cdh1 and the first evidence that Cdh1 may activate the APC by binding specific substrates.
Sørensen
C S
CS
Institute of Cancer Biology, Danish Cancer Society, DK-2100 Copenhagen Ø, Denmark.
Lukas
C
C
Kramer
E R
ER
Peters
J M
JM
Bartek
J
J
Lukas
J
J
D000595:Amino Acid Sequence; D000705:Anaphase; D064173:Anaphase-Promoting Complex-Cyclosome; D000818:Animals; D001665:Binding Sites; D042846:CDC2-CDC28 Kinases; D002453:Cell Cycle; D002478:Cells, Cultured; D017124:Conserved Sequence; D019925:Cyclin A / Q000378:metabolism*; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D016193:G1 Phase; D006801:Humans; D008025:Ligases / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D051381:Rats; D016196:S Phase; D013379:Substrate Specificity; D014407:Tumor Cells, Cultured; D043743:Ubiquitin-Protein Ligase Complexes; D044767:Ubiquitin-Protein Ligases; D014452:Ubiquitins / Q000378:metabolism
D019925:Cyclin A; D014452:Ubiquitins; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome; D044767:Ubiquitin-Protein Ligases; D017346:Protein Serine-Threonine Kinases; D042846:CDC2-CDC28 Kinases; C495894:CDK2 protein, human; C495892:Cdk2 protein, rat; D051357:Cyclin-Dependent Kinase 2; D018844:Cyclin-Dependent Kinases; D008025:Ligases