Core 2: Mammalian Artificial Chromosome (MAC)

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The mammalian artificial chromosome (MAC) Core will support the overall Project, which seeks to address the downstream consequences in cells and animals of DNA damage, the causes and consequences of chromosome missegregation at cell division, and the mechanisms that drive these processes in tumors. It will serve as a foundry for MACs that will support each of the individual projects that require chromosome manipulations and engineering: from site-directed and inducible DNA damage, to reporters for chromosome segregation errors, to housing the genetically encoded components of a chemical genetics toolbox. MACs can carry large (i.e. 100-2000 kb) genetic payloads, and exist in cells as molecules that are propagated and inherited alongside the natural chromosomes. Their autonomous nature is a key advantage, since they deliver their genetic payloads without insertion or other disruption of the host genome. The Core will employ our recent innovations in MAC technology that gave rise to a new generation of MAC templates in which we direct the seeding of centromeric chromatin. This approach permits efficient MAC formation on diverse DNA templates. The Core will undertake MAC approaches from their design, creation, delivery into cells, characterization, through to their implementation for experiments proposed in the individual Projects of the program. Specifically, the Core will develop and use MACs that fall into the following categories: ones that serve as targets for DNA damage, ones that harbor elaborate genetic payloads that will modulate chromosome segregation with a high degree of sophistication, and ones that carry fluorescent reporters of chromosome missegregation. The MAC Core is designed to be flexible to help Project members develop new lines of experimentation that arise from the findings achieved during the initial steps in the overall program.