Elucidating chromosome structure and function through the lens of SMC complexes and R-loops

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The study of budding yeast has provided fundamental insights into the cell biological processes of all eukaryotes. Its study has also made critical contributions to the understanding and treatment of human disorders like diabetes, congenital disabilities, and cancer. My laboratory uses budding yeast to interrogate three areas of cell biology, the higher-order structure of chromosomes, the prevention of chromosome damage and rearrangements, and the mitigation of environmental stress. The basic unit of chromosomes is chromatin, which is composed of the DNA and associated proteins. The organization of chromatin into higher-order structures is essential for high fidelity chromosome segregation, the repair of DNA damage, and the regulation of gene expression. The molecular mechanisms that organize chromatin are major mysteries in cell biology. In this proposal, we study chromatin organization through the analysis of cohesin, a member of the SMC (Structural Maintenance of Chromosomes) family of protein complexes. Cohesin contributes to chromosome organization by tethering together different chromatin regions within a chromosome or between chromosomes. Cohesin also translocates along a chromosome to extrude loops. This proposal interrogates the molecular mechanisms underlying cohesin's tethering and loop-extrusion activities, the regulation of these activities, and their impact on chromosome structure and function in living cells. Cells also maintain genome stability by preventing and repairing chromosome damage. Chromosome damage is often caused by errors in the execution of intrinsic cellular processes. Indeed, during transcription, an RNA transcript can erroneously hybridize with homologous double-stranded DNA sequences on the chromosomes to generates an RNA-DNA hybrid and a displaced single-stranded DNA. This unusual structure, called an R-loop, can cause DNA damage and chromosome rearrangements. Here, we present experiments to understand why only a subset of R-loops in a genome cause DNA damage and how this damage leads to the large chromosome rearrangements that are a common feature of cancer cells. Finally, cellular stress also arises from extrinsic environmental changes. Understanding how some organisms survive extreme environmental changes has provided critical technical and conceptual advances in biology. We study the ability of yeast to survive desiccation. We showed that the expression of a small protein and simple sugar in yeast is necessary and sufficient for yeast to survive desiccation. These two factors prevent the aggregation of model proteins and modulate membranes in vitro. Here, we propose to elucidate the remarkable biological functions of these two factors by identifying the specific cellular proteins and membranes that they protected during desiccation. These studies will provide fundamental insights into protein and membrane homeostasis beyond desiccation and may generate potential novel applications for biomedicine and agriculture.