Dephosphorylation of Cdc20 is required for its C-box-dependent activation of the APC/C.
The anaphase-promoting complex/cyclosome (APC/C) ubiquitin ligase is tightly regulated to ensure programmed proteolysis in cells. The activity of the APC/C is positively controlled by cyclin-dependent kinase (CDK), but a second level of control must also exist because phosphorylation inactivates Cdc20, a mitotic APC/C co-activator. How Cdc20 is dephosphorylated specifically, when CDK is high, has remained unexplained. Here, we show that phosphatases are crucial to activate the APC/C. Cdc20 is phosphorylated at six conserved residues (S50/T64/T68/T79/S114/S165) by CDK in Xenopus egg extracts. When all the threonine residues are phosphorylated, Cdc20 binding to and activation of the APC/C are inhibited. Their dephosphorylation is regulated depending on the sites and protein phosphatase 2A, active in mitosis, is essential to dephosphorylate the threonine residues and activate the APC/C. Consistently, most of the Cdc20 bound to the APC/C in anaphase evades phosphorylation at T79. Furthermore, we show that the 'activation domain' of Cdc20 associates with the Apc6 and Apc8 core subunits. Our data suggest that dephosphorylation of Cdc20 is required for its loading and activation of the APC/C ubiquitin ligase.
Cell Cycle Control Group, UCL Cancer Institute, University College London, London, UK.
Helene
H
Labit
Kazuyuki
K
Fujimitsu
N Sumru
NS
Bayin
Tohru
T
Takaki
Julian
J
Gannon
Hiroyuki
H
Yamano
D064173:Anaphase-Promoting Complex-Cyclosome; D000818:Animals; D064199:Cdc20 Proteins; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D002478:Cells, Cultured; D004789:Enzyme Activation; D005260:Female; D051379:Mice; D008807:Mice, Inbred BALB C; D008938:Mitosis / Q000235:genetics / Q000502:physiology; D008954:Models, Biological; D010749:Phosphoprotein Phosphatases / Q000378:metabolism* / Q000502:physiology; D010766:Phosphorylation; D011485:Protein Binding; D011499:Protein Processing, Post-Translational / Q000235:genetics; D017434:Protein Structure, Tertiary / Q000502:physiology; D018411:Spodoptera; D043743:Ubiquitin-Protein Ligase Complexes / Q000737:chemistry / Q000378:metabolism*; D029867:Xenopus Proteins / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D014982:Xenopus laevis
D064199:Cdc20 Proteins; C577501:Cdc20 protein, Xenopus; D018797:Cell Cycle Proteins; D029867:Xenopus Proteins; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome; D010749:Phosphoprotein Phosphatases