Functional analysis of the MCM2-7 DNA entry/exit gate during S-phase

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The main objective of this research proposal is to uncover how regulated MCM2-7 ring opening affects replicative helicase activation, DNA synthesis and DNA repair, which together will unravel a completely new pathway that is essential for genome stability.Precise DNA replication is an essential hallmark of all living cells. Thus, eukaryotic cells have evolved a sophisticated system to ensure accurate chromosome duplication. By dividing DNA replication into two steps, cells ensure that their genetic material is replicated only once per cell cycle. In the first step, the MCM2-7 (mini chromosome maintenance) hexamer is assembled in a macromolecular ring structure around double-stranded origin DNA, promoting helicase loading onto origins of DNA replication. This complex gets activated during the second step in early S-phase by phosphorylation and the interaction with a number of replication factors, resulting in extrusion of one DNA-strand from its ring, but the molecular details of this step remained elusive.In this project I will use an exclusively available, chemical biology approach allowing the controlled closure of a helicase interface within the MCM2-7 complex, which was previously used to identify the DNA entry gate of MCM2-7.Using state-of-the-art techniques, I want to tackle the following three key aims: 1) Identification of the DNA exit gate of MCM2-7; 2) Characterisation of the protein complexes that activate the replicative helicase; and 3) Comprehension of potential MCM2-7 ring opening requirements during DNA synthesis in the absence and presence of DNA damage. Funding of this pioneering and interdisciplinary research proposal by the DFG will allow me to dissect a completely novel mechanism in DNA replication, the regulated MCM2-7 ring opening. This work will have major relevance for multiple aspects of DNA replication and DNA repair. In addition, the project may also form the basis for future chemotherapeutics development with anti-cancer activity, as the inhibition of MCM2-7 ring opening offers an efficient route to block proliferation of cancer cells.