All eukaryotic cells depend on faithful cell division to give rise to daughter cells that contain the correct number of chromosomes. Chromosome segregation errors during meiosis in germ cells can lead to aneuploid gametes, which can cause infertility, genetic disorders such as Down Syndrome and miscarriages. Additionally, chromosome segregation errors during mitosis can lead to aneuploid somatic cells, which are a hallmark of cancer. Despite the importance of tightly regulating these two types of cell division, the mechanisms that ensure fidelity during these processes remain active areas of investigation. Proteins that have been shown to be important regulators of fidelity during meiosis in Caenorhabditis elegans (C. elegans) include ZHP-3 and PCH-2. ZHP-3 is a putative SUMO or ubiquitin ligase required for crossover recombination, but whether or not ZHP-3 is regulated to promote recombination remains poorly understood. PCH-2 is a conserved AAA+ ATPase required for timely coordination of the meiotic prophase events pairing, synapsis and recombination, but the mechanism for how it accomplishes this coordination is currently unknown. Additionally, PCH-2 has been shown to be required for both silencing and activation of the spindle assembly checkpoint during mitosis, but how it regulates the checkpoint is still an active area of investigation. Here, I use C. elegans as a model system to investigate the molecular mechanisms of these two proteins during meiotic and mitotic chromosome segregation. In Chapter 1, I investigate whether ZHP-3 is phosphorylated by a conserved DNA damage kinase, CHK-1, to determine if this phosphorylation event is required for ZHP-3’s localization or function during meiosis. I find that while ZHP-3 is phosphorylated by CHK-1 kinase in vitro, mutation of these phosphorylation sites in vivo does not affect ZHP-3’s localization to meiotic chromosomes, or its function in promoting crossover recombination. In Chapter 2, I investigate a different regulator of meiosis, PCH-2, to determine how it coordinates meiotic prophase events and whether it acts on meiotic HORMADS to accomplish this. I find that mutations in PCH-2 genetically interact with mutations in the meiotic HORMADS HTP-3 and HIM-3, suggesting that these proteins work together to coordinate meiotic prophase events. Additionally, I find that PCH-2 acts on HTP-3 to coordinate pairing and synapsis, but acts on HIM-3 during recombination, indicating that the meiotic HORMADS are cleanly delegated to regulate specific meiotic prophase events. Finally, in Chapter 3, I explore PCH-2’s role during somatic cell division and investigate how PCH-2 regulates the strength of the spindle assembly checkpoint. I find that PCH-2 is more enriched in P1 cells compared to AB cells in the early C. elegans embryo, potentially explaining why cells that give rise to the germline have stronger checkpoints than somatic cells. Additionally, we find that this enrichment depends on cell fate factors and PCH-2’s adaptor protein CMT-1. Taken together, these results contribute mechanistic insight for how these proteins promote fidelity during meiosis and mitosis by determining post-translational modifications of meiotic recombination factors (Chapter 1), identifying potential substrates of meiotic prophase regulators (Chapter 2), and demonstrating how spindle checkpoint regulators are differentially inherited in specific cell types to give rise to differences in spindle checkpoint strength (Chapter 3).
