Identification and characterisation of Ubiquitin Specific Protease 11 binding partners

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Title: Identification and characterisation of Ubiquitin Specific Protease 11 binding partners
Authors: Stockum, Anna
Item Type: Thesis or dissertation
Abstract: Throughout the last decades, post-translational modification of proteins by ubiquitination has proved to be important for most cellular processes. The regulation depends on ubiquitin addition, which is orchestrated by a cascade of three enzymes, as well as ubiquitin removal, catalysed by deubiquitinating enzymes (DUBs). The human genome encodes nearly 100 DUBs of which the Ubiquitin Specific Proteases (USPs) are the largest of the five DUB families. This project focuses on USP11, which was previously thought to play a role in DNA damage response (DDR). In order to further understand the possible role for USP11 in DDR, we have identified several USP11 interacting proteins, namely PAM, SPRYD3 and RAE1. We show that SPRYD3 and RAE1 are ubiquitinated and also identify these proteins as USP11 substrates. The E3 ligase, PAM functions as a SCF-like complex with the F-box protein FBXO45. Our data shows strong similarities between this interaction and the interaction of PAM with SPRYD3, suggesting that the two proteins could form an alternative E3 ligase complex. Another USP11 interacting protein, RAE1, also interacts with PAM, which was verified in this study. RAE1 was identified as an RNA export factor, but was recently shown to be involved in regulation of the mitotic checkpoint in a complex with NUP98. Indeed, growth curves of RAE1 depleted U2OS cells show a significant lack in cell proliferation compared to control shRNA expressing cells. Surprisingly, a similar growth defect was observed in USP11 depleted cells. Ablation of RAE1 or USP11 allowed a small fraction of nocodazole arrested cells to proceed in the cell cycle, suggesting that the mitotic checkpoint is affected by the lack of either of these proteins. While further investigations are required to prove our hypotheses, the presented work identifies a variety of new substrates for USP11 and a new pathway it appears to regulate.
Content Version: Open Access
Issue Date: Mar-2015
Date Awarded: Jul-2015
URI: http://hdl.handle.net/10044/1/65757
Supervisor: Maertens, Goedele
McClure, Myra
Sponsor/Funder: Imperial College London
Department: Department of Medicine
Publisher: Imperial College London
Qualification Level: Doctoral
Qualification Name: Doctor of Philosophy (PhD)
Appears in Collections:Medicine PhD theses



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