Discovery and Elucidation of Novel Regulators of Cell Division

Abstract

Mitotic cell division is a process requiring a highly coordinated dance between many enzymes, substrates and metabolites to result in the segregation of identical sets of daughter chromosomes. A hallmark of cancer is the ability to perturb this process in ways that increase the proliferation of cancer cells. We have studied several aspects of cellular division in order to further elucidate how cancer cell progression can occur in human disease. Mammalian cell division is a biological process that has been studied for decades and important discoveries often coincide with the development of noveltools and techniques. I have developed a new cell-based high-throughput screening tool that combines CRISPR/Cas9 technology with the Fluorescence Ubiquitin Cell Cycle Indicator system (FUCCI) for assessing the cell cycle effects of knocking out genes of interest. This tool provides a genome-encoded cell cycle phase indicator system and dox-inducible Cas9 for use with guide RNA libraries for future screens. Many of the proteins our lab studies were initially identified through either proteomic analysis or genomic screening. While these approaches have yielded interesting hits, they have focused exclusively on protein-based regulation of cell cycle progression and division. To this end we performed a high-throughput screen of 1,200 different naturally occurring metabolites in order to find novel affectors of the cell cycle and have identified 180 putative. These results will provide the basis for future projects analyzing these metabolites and their roles in cell cycle regulation. Ribosome biogenesis has long been linked to cell proliferation and in my studies, I characterized Rexo4, an exonuclease responsible for processing nascent ribosomal RNA. Recent studies suggest that Rexo4 is a biomarker for cancer disease and is seen to be upregulated in cancer cells at both the mRNA and protein level. My work has determined that Rexo4 is a requirement for cell cycle progression in mammalian cells and that both its nucleolar localization and exonuclease activity are required for cell proliferation