Investigating the Role of Orcs in Chromatin Organization and in Mitotic Progression

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This award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5).Intellectual merit: How the accurate duplication of genetic material and its faithful segregation to daughter cells is coordinated in multi-cellular organisms is an important question that remains to be clearly understood. In eukaryotic cells these two events occur in alternative oscillating cycles. Initiation of DNA replication requires the ordered assembly of a multiprotein complex including the Origin Recognition Complex (ORC) at origins of DNA replication. In mammalian cells chromosomal domains occupy specific nuclear positions and replicate at defined times during S-phase of the cell division cycle. Chromatin changes and specific modifications orchestrate the establishment of spatio-temporal profiling of DNA replication that is ORC-mediated. ORC proteins are critical not only for initiation of DNA replication, but have also been implicated in non-preRC functions including heterochromatin organization and silencing, chromosome segregation and mitosis in several model organisms. The objective of this project is to address how ORCs coordinate DNA replication with chromatin organization and mitotic progression in mammalian cells. The association of ORC subunits with replication origins, heterochromatin, centromeres/kinetochores, centrosomes and cytokinetic furrow suggests that ORCs play a pivotal role in coordinating all stages of chromosome inheritance cycle by interacting with different protein partners in a cell-cycle dependent fashion. This project will identify the roles played by novel protein partners that bind to ORCs during different stages of the cell cycle that will address how ORC participates in various cellular processes. In particular this project will provide crucial insights into the role of ORCs in the recruitment and spreading of chromatin proteins to heterochromatin. Finally, several approaches will be used to understand whether the role of ORCs in chromatin organization and mitosis is independent of its function in DNA replication initiation. Since the involvement of ORCs in chromatin organization is conserved from yeast to humans the outcome of this project will have broad scientific impact.Broader impacts: This research is amenable to an active outreach program since it utilizes extensive microscopy and live cell imaging that is visually appealing and brings out the power of cell biology in understanding basic concepts of Biology. Being in a University setup gives a unique opportunity to mentor undergraduate, graduate students and postdoctoral fellows. This project integrates the use of Biochemistry, Molecular Biology and Cell Biology to capture the interest of students not only from Biology background but also from Chemistry and Bioengineering. This project will also involve providing students with a direct view of the concepts cell and molecular biology including actively mentoring undergraduate students, including women and minorities. This project will enable the understanding of a basic biology question, on how a cell accomplishes DNA replication and signals it to undergo chromosome segregation and cell division.