The deubiquitylase USP15 regulates topoisomerase II alpha to maintain genome integrity

Andrew B Fielding,Ian A Prior,Michael J Clague,Sylvie Urbé,Matthew Concannon,Judy M Coulson,Sarah Darling,Joseph J Sacco,Emma V Rusilowicz-Jones

doi:10.1038/s41388-017-0092-0

Andrew B Fielding, Ian A Prior + Show 7 more

Open Access

https://doi.org/10.1038/s41388-017-0092-0

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Journal: Oncogene	Publication Date: Feb 12, 2018
Citations: 30	License type: open-access

Affiliation: University of Liverpool

Abstract

Ubiquitin-specific protease 15 (USP15) is a widely expressed deubiquitylase that has been implicated in diverse cellular processes in cancer. Here we identify topoisomerase II (TOP2A) as a novel protein that is regulated by USP15. TOP2A accumulates during G2 and functions to decatenate intertwined sister chromatids at prophase, ensuring the replicated genome can be accurately divided into daughter cells at anaphase. We show that USP15 is required for TOP2A accumulation, and that USP15 depletion leads to the formation of anaphase chromosome bridges. These bridges fail to decatenate, and at mitotic exit form micronuclei that are indicative of genome instability. We also describe the cell cycle-dependent behaviour for two major isoforms of USP15, which differ by a short serine-rich insertion that is retained in isoform-1 but not in isoform-2. Although USP15 is predominantly cytoplasmic in interphase, we show that both isoforms move into the nucleus at prophase, but that isoform-1 is phosphorylated on its unique S229 residue at mitotic entry. The micronuclei phenotype we observe on USP15 depletion can be rescued by either USP15 isoform and requires USP15 catalytic activity. Importantly, however, an S229D phospho-mimetic mutant of USP15 isoform-1 cannot rescue either the micronuclei phenotype, or accumulation of TOP2A. Thus, S229 phosphorylation selectively abrogates this role of USP15 in maintaining genome integrity in an isoform-specific manner. Finally, we show that USP15 isoform-1 is preferentially upregulated in a panel of non-small cell lung cancer cell lines, and propose that isoform imbalance may contribute to genome instability in cancer. Our data provide the first example of isoform-specific deubiquitylase phospho-regulation and reveal a novel role for USP15 in guarding genome integrity.

Highlights

Ubiquitylation is a reversible post-translational modification that can target proteins for degradation or regulate their activity or cellular localisation [1]
We found that A549 lung adenocarcinoma cells express both ubiquitin-specific protease 15 (USP15) isoforms, with higher levels of isoform-1, which can be distinguished by isoform-specific small interfering RNAs and reverse transcription-PCR (RT-PCR) primers (Supplementary Figure S1)
We identified TOP2A as a novel protein that was consistently downregulated by a pool of USP15 small interfering RNAs (siRNAs) (Fig. 1a)

Summary

Introduction

Ubiquitylation is a reversible post-translational modification that can target proteins for degradation or regulate their activity or cellular localisation [1]. DUBs can be regulated by conformational changes, adaptor proteins, or posttranslational modifications, which control their activity or recruitment to specific complexes [11, 12]. During the cell cycle, periodic phosphorylation activates USP16 and USP37 [14, 15] but inactivates USP8 through recruitment of 14-3-3 proteins [16]. The regulated expression of DUBs may control their cellular availability, and alternative splicing can generate DUB isoforms that are targeted to distinct subcellular compartments, as described for USP33 [17], or exhibit different substrate specificity, as recently suggested for ubiquitin-specific protease 15 (USP15) [18]

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