Abstract
The anaphase‐promoting complex/cyclosome (APC/C), a multi‐subunit ubiquitin ligase essential for cell cycle control, is regulated by reversible phosphorylation. APC/C phosphorylation by cyclin‐dependent kinase 1 (Cdk1) promotes Cdc20 co‐activator loading in mitosis to form active APC/C‐Cdc20. However, detailed phospho‐regulation of APC/C dynamics through other kinases and phosphatases is still poorly understood. Here, we show that an interplay between polo‐like kinase (Plx1) and PP2A‐B56 phosphatase on a flexible loop domain of the subunit Apc1 (Apc1‐loop500) controls APC/C activity and mitotic progression. Plx1 directly binds to the Apc1‐loop500 in a phosphorylation‐dependent manner and promotes the formation of APC/C‐Cdc20 via Apc3 phosphorylation. Upon phosphorylation of loop residue T532, PP2A‐B56 is recruited to the Apc1‐loop500 and differentially promotes dissociation of Plx1 and PP2A‐B56 through dephosphorylation of Plx1‐binding sites. Stable Plx1 binding, which prevents PP2A‐B56 recruitment, prematurely activates the APC/C and delays APC/C dephosphorylation during mitotic exit. Furthermore, the phosphorylation status of the Apc1‐loop500 is controlled by distant Apc3‐loop phosphorylation. Our study suggests that phosphorylation‐dependent feedback regulation through flexible loop domains within a macromolecular complex coordinates the activity and dynamics of the APC/C during the cell cycle.
Highlights
Cells use a highly conserved ordered mechanism, cell cycle control, to ensure that complete genomes are accurately duplicated and transmitted to daughter cells
We found that a flexible loop domain of Apc1 [Apc1-loop500: 515– 584 in Xenopus] contains two tandem copies of the polo-binding motifs “S-[pS/pT]-[P/X]” (PBMs), in which X indicates any amino acid
When wild-type (WT) Apc1-loop500 fragment fused to maltose-binding protein (MBP) was incubated with anaphase extracts supplemented with non-degradable cyclin B, to ensure continuation of the anaphase state even after activation of the anaphase-promoting complex/cyclosome (APC/C), endogenous Plx1 binding to Apc1-loop500 was observed, over and above binding to MBP itself (Fig 1B)
Summary
Cells use a highly conserved ordered mechanism, cell cycle control, to ensure that complete genomes are accurately duplicated and transmitted to daughter cells. Cdk1-cyclin B complex phosphorylates and activates the anaphase-promoting complex/cyclosome (APC/C) E3 ubiquitin ligase (Morgan, 2007; Pines, 2011; Primorac & Musacchio, 2013; Alfieri et al, 2017). The activated APC/C triggers the transition from metaphase to anaphase by targeting securin and cyclin B for proteasome-dependent degradation, which results in chromosome separation and inactivation of Cdk, respectively. The phosphorylation states of the APC/C dynamically change in a manner coordinated with the progression of mitosis. It is still poorly understood how phosphorylation of APC/C is precisely regulated during mitosis, a key element of the working of the APC/ C system
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