Chromosome structure, duplication and stability in yeast

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Project Summary/Abstract Eukaryotic genome duplication requires that each chromosomal base pair is copied efficiently, accurately and only once per cell division, a monumental demand given the millions to billions of base pairs that comprise eukaryotic genomes and the countless cell divisions required to form and sustain organisms. Severe defects in DNA replication are incompatible with life. However, mild perturbations in this process, while still capable of supporting cell division, and which can be challenging to identify using biochemical approaches, can compromise development and health over the course of multiple cell divisions. Regulation of the first step, the initiation of DNA replication that occurs at chromosomal positions called origins, is particularly critical in eukaryotic cells because their chromosomes require multiple spatially and temporally distributed origins for accurate and efficient duplication. Perturbations in origin number or distribution can promote cancer, stem cell aging, or developmental disorders. While the origin-binding proteins and molecular steps that define an origin are known, the mechanisms that regulate chromosomal origin number and distribution are unclear. A challenge is that chromatin heterogeneity exists across chromosomes as an intrinsic part of genome functional organization. Thus, the origin-binding proteins must work sufficiently enough within distinct chromatin environments to achieve a level of origin distribution that balances the competing demands for cell proliferation and genome stability. Dr. Fox's lab addresses the gaps in understanding how native chromatin structures regulate origin function by combining rigorous genetics and genomics to reveal chromatin-mediated mechanisms that impinge on the structure and function of Saccharomyces cerevisiae (yeast) origins. Emphasis is placed on the first step of origin formation, the origin licensing reaction, which occurs in G1-phase of the cell cycle that precedes the S-phase where origins perform their actual function, unwinding of the parental DNA for new DNA synthesis. Accumulating evidence reveals that the licensing step is particularly relevant to both genome stability and cell-fate decisions, but there is a paucity of molecular mechanisms regarding how it is regulated in vivo to achieve chromosomal origin distribution. Evolutionary conservation of the origin-binding proteins and multiple features of chromatin allow the Fox laboratory to leverage the experimental strengths of yeast to define fundamental mechanisms by which chromatin and the origin-binding proteins collaborate to form and distribute origins over the genome.