PD01-01 VHL-MEDIATED UBIQUITINATION OF THE KINASE MPS1 REGULATES THE MITOTIC CHECKPOINT IN CLEAR CELL RENAL CELL CARCINOMA

Abstract

You have accessJournal of UrologyCME1 May 2022PD01-01 VHL-MEDIATED UBIQUITINATION OF THE KINASE MPS1 REGULATES THE MITOTIC CHECKPOINT IN CLEAR CELL RENAL CELL CARCINOMA Mark Woodford, Sarah Backe, Rebecca Sager, Oleg Shapiro, Imad Nsouli, Laura Wengert, Gennady Bratslavsky, Dimitra Bourboulia, and Mehdi Mollapour Mark WoodfordMark Woodford More articles by this author , Sarah BackeSarah Backe More articles by this author , Rebecca SagerRebecca Sager More articles by this author , Oleg ShapiroOleg Shapiro More articles by this author , Imad NsouliImad Nsouli More articles by this author , Laura WengertLaura Wengert More articles by this author , Gennady BratslavskyGennady Bratslavsky More articles by this author , Dimitra BourbouliaDimitra Bourboulia More articles by this author , and Mehdi MollapourMehdi Mollapour More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000002516.01AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Abnormal chromosome segregation during mitosis causes aneuploidy, a hallmark of cancers associated with high risk for tumorigenesis, which is normally regulated by the mitotic checkpoint. Mps1 kinase activity is essential for spindle checkpoint signaling. Mps1 is over-expressed in clear cell renal cell carcinoma (ccRCC) and requires the molecular chaperone Hsp90 for its activity. Mps1 phosphorylates Hsp90, regulating chaperone function of numerous oncogenic client proteins, including Mps1, and conferring tumor selectivity of Hsp90 inhibitors in ccRCC. The most frequent alteration leading to ccRCC is loss of von Hippel Lindau (VHL), the recognition subunit of an E3-ubiquitin ligase complex that targets proteins for degradation. As Mps1 is over-expressed in ccRCC, the objective of the current study was to determine whether Mps1 gets targeted for degradation by VHL and whether this regulates the mitotic checkpoint. METHODS: VHL-mediated ubiquitination of Mps1 was examined in vitro and VHL-dependent degradation of Mps1 was examined in ccRCC cell lines by Western blot. Site-directed mutagenesis was utilized to determine Mps1 ubiquitination sites and the effect of increased Mps1 stability on mitotic checkpoint was determined using flow cytometry. RESULTS: Mps1 kinase is ubiquitinated by a VHL containing E3-ubiquitin ligase complex and Mps1 activity is essential for its ubiquitination. Re-expression of VHL in VHL-null ccRCC cell lines leads to proteasomal degradation of Mps1. VHL degrades Mps1 in an oxygen independent manner by ubiquitination of Mps1-K86, K827, and K848. Mps1 ubiquitination regulates cell cycle progression via exit from the mitotic checkpoint. CONCLUSIONS: Mps1 is targeted for degradation by the tumor suppressor VHL in a hypoxia-independent manner and Mps1 is over-expressed in VHL-null ccRCC. VHL-mediated ubiquitination of Mps1 regulates mitotic checkpoint progression. Mps1 stability additionally mediates Hsp90 post-translational modification and tumor selectivity of Hsp90 inhibitors. Source of Funding: This work was also supported by the National Institute of General Medical Sciences of the National Institutes of Health under Award Number, R01GM139932 (D.B.), R35GM139584 (M.M.), and R01GM124256 (M.M.). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. This work was also supported by the Urology Care Foundation-American Urological Association (M.M.) © 2022 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 207Issue Supplement 5May 2022Page: e30 Advertisement Copyright & Permissions© 2022 by American Urological Association Education and Research, Inc.MetricsAuthor Information Mark Woodford More articles by this author Sarah Backe More articles by this author Rebecca Sager More articles by this author Oleg Shapiro More articles by this author Imad Nsouli More articles by this author Laura Wengert More articles by this author Gennady Bratslavsky More articles by this author Dimitra Bourboulia More articles by this author Mehdi Mollapour More articles by this author Expand All Advertisement PDF DownloadLoading ...