The Spindle Assembly Checkpoint Is Not Essential for Viability of Human Cells with Genetically Lowered APC/C Activity.

Thomas Wild,Takeo Narita,Julie Schou,Jakob Nilsson,Chunaram Choudhary,Marie Sofie Yoo Larsen

doi:10.1016/j.celrep.2016.01.060

Thomas Wild, Takeo Narita + Show 4 more

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https://doi.org/10.1016/j.celrep.2016.01.060

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Journal: Cell Reports	Publication Date: Feb 18, 2016
Citations: 49	License type: cc-by

Affiliation: Novo Nordisk Foundation, University of Copenhagen

Abstract

SummaryThe anaphase-promoting complex/cyclosome (APC/C) and the spindle assembly checkpoint (SAC), which inhibits the APC/C, are essential determinants of mitotic timing and faithful division of genetic material. Activation of the APC/C is known to depend on two APC/C-interacting E2 ubiquitin-conjugating enzymes—UBE2C and UBE2S. We show that APC/C activity in human cells is tuned by the combinatorial use of three E2s, namely UBE2C, UBE2S, and UBE2D. Genetic deletion of UBE2C and UBE2S, individually or in combination, leads to discriminative reduction in APC/C function and sensitizes cells to UBE2D depletion. Reduction of APC/C activity results in loss of switch-like metaphase-to-anaphase transition and, strikingly, renders cells insensitive to chemical inhibition of MPS1 and genetic ablation of MAD2, both of which are essential for the SAC. These results provide insights into the regulation of APC/C activity and demonstrate that the essentiality of the SAC is imposed by the strength of the APC/C.

Highlights

The anaphase-promoting complex/cyclosome (APC/C) is a multi-subunit E3 ubiquitin ligase that is essential for eukaryotic cell division (Peters, 2006; Pines, 2011; Primorac and Musacchio, 2013; Sivakumar and Gorbsky, 2015)
We show that anaphase-promoting complex/cyclosome (APC/ C) activity in human cells is tuned by the combinatorial use of three E2s, namely UBE2C, UBE2S, and UBE2D
Reduction of APC/C activity results in loss of switch-like metaphase-toanaphase transition and, strikingly, renders cells insensitive to chemical inhibition of monopolar spindle 1 (MPS1) and genetic ablation of MAD2, both of which are essential for the spindle assembly checkpoint (SAC)

Summary

Introduction

The anaphase-promoting complex/cyclosome (APC/C) is a multi-subunit E3 ubiquitin ligase that is essential for eukaryotic cell division (Peters, 2006; Pines, 2011; Primorac and Musacchio, 2013; Sivakumar and Gorbsky, 2015). The APC/C promotes ubiquitylation-mediated degradation of key mitotic regulators, such as securin and cyclin B1 (CCNB1), which is required for metaphase-to-anaphase transition and cell division. The APC/C employs two E2 ubiquitin-conjugating enzymes, UBE2C (UBCH10) and UBE2S, in tandem, whereby UBE2C initiates substrate ubiquitylation and UBE2S subsequently extends substrate-linked ubiquitin to exclusively generate K11-linked polyubiquitin chains (Garnett et al, 2009; Summers et al, 2008; Williamson et al, 2009; Wu et al, 2010). The timing of APC/C activation in mitosis is tightly controlled by the spindle assembly checkpoint (SAC) (London and Biggins, 2014), which senses kinetochores that are not attached to microtubules and generates the mitotic checkpoint complex (MCC). Inactivation of the SAC in mammalian cells invariably leads to catastrophic aneuploidy, and, genetic deletions of SAC components are lethal (Kops et al, 2005)

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