Complete and accurate DNA replication is essential to maintain the genetic integrity in all organisms. In eukaryotes, the minichromosome maintenance (MCM) complex forms the catalytic core of the CMG helicase that unwinds DNA at the replication fork. We have previously identified a conserved MCM complex binding protein (MCM-BP) through a proteomic screen in human cells. In chapter two of this thesis, I show that MCM-BP makes an important contribution to nuclear morphology in human cells by affecting centrosome duplication. I also show that MCM-BP depletion results in G2 checkpoint signaling and the induction of replication stress. A recent study in Xenopus egg extracts has suggested that MCM-BP functions to unload the MCM complex from chromatin during S-phase. However, the mechanism of this process remains enigmatic. In chapter three of this thesis, I show that MCM-BP directly binds the de-ubiquitylating enzyme, USP7 and that this interaction is mediated by S158 on MCM-BP and the USP7 TRAF domain. Furthermore, I indicate a novel role for USP7 in DNA replication that involves unloading of the MCM complex during S-phase. Finally, my data suggest that MCM-BP tethers an interaction between the USP7 and the MCM complex to facilitate MCM complex unloading at the end of S-phase.
| Identifer | oai:union.ndltd.org:TORONTO/oai:tspace.library.utoronto.ca:1807/65669 |
| Date | 21 July 2014 |
| Creators | Jagannathan, Madhav |
| Contributors | Frappier, Lori |
| Source Sets | University of Toronto |
| Language | en_ca |
| Detected Language | English |
| Type | Thesis |
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