53BP1 and USP28 mediate p53-dependent cell cycle arrest in response to centrosome loss and prolonged mitosis.

Chii Shyang Fong,Gregory Mazo,Meng-Fu Bryan Tsou,Joshua Goodman,Brian P O'Rourke,Denisse Izquierdo,Minhee Kim,Tuhin Das

doi:10.7554/elife.16270

Chii Shyang Fong, Gregory Mazo + Show 6 more

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https://doi.org/10.7554/elife.16270

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Abstract

Mitosis occurs efficiently, but when it is disturbed or delayed, p53-dependent cell death or senescence is often triggered after mitotic exit. To characterize this process, we conducted CRISPR-mediated loss-of-function screens using a cell-based assay in which mitosis is consistently disturbed by centrosome loss. We identified 53BP1 and USP28 as essential components acting upstream of p53, evoking p21-dependent cell cycle arrest in response not only to centrosome loss, but also to other distinct defects causing prolonged mitosis. Intriguingly, 53BP1 mediates p53 activation independently of its DNA repair activity, but requiring its interacting protein USP28 that can directly deubiquitinate p53 in vitro and ectopically stabilize p53 in vivo. Moreover, 53BP1 can transduce prolonged mitosis to cell cycle arrest independently of the spindle assembly checkpoint (SAC), suggesting that while SAC protects mitotic accuracy by slowing down mitosis, 53BP1 and USP28 function in parallel to select against disturbed or delayed mitosis, promoting mitotic efficiency.

Highlights

Mitosis is a critical cell cycle phase during which duplicated chromosomes are correctly separated into two identical units on the spindle, restoring genome integrity after cell division
Non-transformed retinal pigment epithelial (RPE) cells, we constructed a stable PLK4as cell line in which the endogenous PLK4, a kinase required for centrosome duplication (Habedanck et al, 2005; Bettencourt-Dias et al, 2005), was replaced with an analog-sensitive mutant (PLK4as) that could be chemically inactivated by the ATP analog 3MBPP1 (Kim, 2016)
While the accuracy of chromosome segregation is guarded by spindle assembly checkpoint (SAC), we found that a separate program involving 53BP1 and USP28 is required to monitor mitotic progression independent of SAC

Summary

Introduction

Mitosis is a critical cell cycle phase during which duplicated chromosomes are correctly separated into two identical units on the spindle, restoring genome integrity after cell division. After committing to mitotic entry, the accuracy of chromosome segregation is further protected by the spindle assembly checkpoint (SAC), which functions to delay anaphase onset until all chromosomes or kinetochores are properly attached to spindle microtubules (Rieder and Cole, 2000; LaraGonzalez et al, 2012). A wide range of cellular stresses are known to directly or indirectly disturb spindle assembly or chromosome segregation, including centrosome/kinetochore/microtubule dysfunctions (Vitre and Cleveland, 2012; Bazzi and Anderson, 2014), DNA damages (Mikhailov et al, 2002; Carlson, 1950; Smits et al, 2000; Hut et al, 2003), heat shock (Maldonado-Codina et al, 1993; Vidair et al, 1993; Zajac et al, 2008; Erenpreisa et al, 2000), hypoxia (Fischer et al, 2004; Nystul et al, 2003), and oxidative stress (Kurata, 2000), many of which activate SAC to slow down mitosis so that extra time is available for error correction. A missegregating chromosome in anaphase is able to induce p53-dependent cell cycle arrest in the G1

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