PP1 promotes cyclin B destruction and the metaphase–anaphase transition by dephosphorylating CDC20

Abstract

Ubiquitin-dependent proteolysis of cyclin B and securin initiates sister chromatid segregation and anaphase. The anaphase promoting complex/cyclosome and its co-activator CDC20 (APC/CCDC20) form the main ubiquitin E3 ligase for these two proteins. APC/CCDC20 is regulated by CDK1-cyclin B and counteracting PP1 and PP2A family phosphatases through modulation of both activating and inhibitory phosphorylation. Here, we report that PP1 promotes cyclin B destruction at the onset of anaphase by removing specific inhibitory phosphorylation in the N-terminus of CDC20. Depletion or chemical inhibition of PP1 stabilises cyclin B and results in a pronounced delay at the metaphase-to-anaphase transition after chromosome alignment. This requirement for PP1 is lost in cells expressing CDK1-phosphorylation defective CDC206A mutants. These CDC206A cells show a normal spindle checkpoint response, and rapidly destroy cyclin B once all chromosomes have aligned and enter into anaphase in the absence of PP1 activity. PP1 therefore facilitates the metaphase-to-anaphase by promoting APC/CCDC20-dependent destruction of cyclin B in human cells.

Deprotection of centromeric cohesin at meiosis II requires APC/C activity but not kinetochore tension

“…Our data favor the latter possibility because lack of tension at meiosis II delays all APC/C Cdc20 ‐dependent processes in cells lacking MCCs. Work in worms, together with data from yeast and human cells, suggests that the Bub1–Bub3 complex recruits Cdc20 to the kinetochore protein KNL1 where it is exposed to two forms of tension‐sensitive regulation: in the absence of tension, Cdc20 is incorporated into the MCC, whereas in the presence of tension, Cdc20 is dephosphorylated by PP1, which enhances its affinity for the APC/C (Yang et al, 2015; Kim et al, 2017; Bancroft et al, 2020). Thus, APC/C Cdc20 activity is still responsive to tension in cells lacking MCCs.…”

M-Phase oscillations in PP1 activity are essential for accurate progression through mammalian oocyte meiosis

“…Active pools of PP1 in prometaphase impact M-Phase exit and oocyte quality In mitosis, PP1 plays a pivotal role in the metaphase/anaphase transition (e.g., silencing the spindle assembly checkpoint and APC/C CDC20 activation) and cytokinesis. In human mitotic cells, PP1 inhibition or siRNA-mediated loss of PP1 leads to delayed metaphase/anaphase transition, metaphase arrest, and binucleated cells (a sign of cytokinesis failure) (Cheng et al, 2000;Winkler et al, 2015;Bhowmick et al, 2019;Capalbo et al, 2019;Bancroft et al, 2020). We found a similar phenotype in TMC-treated oocytes, with PP1 inhibition for the duration of meiosis (0-16 h post-meiotic arrest release) causing metaphase I arrest or meiotic abnormalities (e.g., chromosome misalignment at metaphase II or cytokinetic failure (two spindles without a PB)).…”

“…PP1mediated dephosphorylation prevents proteasome-mediated degradation of specific proteins (PER2 and MYC) (Gallego et al, 2006;Dingar et al, 2018). Additionally, PP1-mediated dephosphorylation of CDC20 plays a role in activating APC/C CDC20 , the major E3 ligase involved in the degradation of pro-M-Phase proteins at metaphase/anaphase transition (Kim et al, 2017;Bancroft et al, 2020). Degradation of the APC/C CDC20 substrates securin and cyclin B starts in prometaphase of oocytes and is essential for metaphase I/anaphase I transition and accurate chromosome segregation (Levasseur et al, 2019;Thomas et al, 2021).…”

“…PP1 and PP2A are critically important for mitotic and meiotic M-Phase progression and exit. In mitosis, PP1 and PP2A have unique temporal and spatial regulation, and work in concert to oppose CDK1 activity, preventing M-Phase entry and promoting metaphase-to-anaphase transition (Grallert et al, 2015;Ma et al, 2016;Rogers et al, 2016;Bancroft et al, 2020). In mammalian oocytes, inhibition of PP1/PP2A with reagents such as okadaic acid and Calyculin A causes major meiotic abnormalities, and oocyte-specific conditional loss of PP2A causes substantial defects in chromosome segregation and M-Phase progression (Alexandre et al, 1991;Gavin et al, 1991;Schwartz and Schultz, 1991;Smith et al, 1998;Mailhes et al, 2003;Swain et al, 2003;Wang et al, 2004;Swain et al, 2007;Hu et al, 2014;Tang et al, 2016).…”

“…In mammals, there are three PP1c genes and hundreds of RIPPOs, resulting in hundreds of unique holoenzymes with distinctive functions and substrates (Bollen et al, 2010;Heroes et al, 2013). Various forms of PP1 holoenzymes allow PP1 activity to contribute to numerous processes in mitosis, including the G2/M transition, microtubule-kinetochore attachments, the spindle assembly checkpoint, and aspects of cytokinesis (Cheng et al, 2000;Margolis et al, 2003;Winkler et al, 2015;Bhowmick et al, 2019;Capalbo et al, 2019;Conti et al, 2019;Smith et al, 2019;Bancroft et al, 2020). Oscillations in phosphorylation of RIPPOs and PP1c, particularly phosphorylation of PP1c's inhibitory site (pT320 on PP1cα and the equivalent sites in PP1cβ and PP1cγ), during M-Phase play a critical role in the formation of PP1 holoenzymes and PP1 activity, and ultimately M-Phase progression (Dohadwala et al, 1994;Kwon et al, 1997;Nasa et al, 2018).…”