Identification and characterization of regulators of mitotic division and spindle position in Caenorhabditis elegans

The cell division cycle is a highly regulated set of events that results in the generation of two genetically identical cells during M phase. M phase is highly regulated to ensure the fidelity of genome segregation and proper position of the cell division plane. To understand how mitosis and division plane position are regulated, I have studied essential mitotic regulators in Caenorhabditis elegans using several approaches.; The cyclin/Cdk1 complex is the key regulator of entry into and progression through mitosis. To determine the specific roles of cyclin B and Cdk1, we determined the functions of the C. elegans homologs using forward and reverse genetics. C. elegans cdk-1/ncc-1 is absolutely required for meiosis and mitosis and requires the cyb-1, cyb-2.1, cyb-2.2 and cyb-3 cyclin B homologs. These different cyclin B homologs have overlapping and distinct roles in mitosis and localize differently. Thus, different cyclin B/Cdk1 complexes may regulate mitosis through overlapping and distinct sets of effectors.; During mitosis, the mitotic spindle is responsible for chromosome segregation and positionally determines the plane of cell division. To understand how this is accomplished, we have studied the essential spindle component LIN-5. lin-5 was previously shown to be required for chromosome segregation and spindle positioning. We show that LIN-5 localizes the GPR-1 and GPR-2 (G&barbelow; p&barbelow;rotein r&barbelow;egulator) proteins to the spindle and cell cortex, and this complex is required for embryonic spindle positioning and cell division in C. elegans. The GPRs contain a GoLoco motif which can bind and regulate Galpha subunits of heterotrimeric G proteins, thus implicating a G protein signal in spindle function. Two Galpha genes, goa-1 and gpa-16, are required for spindle positioning and cell division, similar to gpr-1,2 and lin-5. Additionally, GOA-1 colocalizes with LIN-5/GPR at the cell cortex and directly binds GPR-1. Activation of Galpha may also require RIC-8, a candidate nucleotide exchange factor for Galpha. We show that ric-8 is required for spindle positioning and cell division, genetically interacts with both Galpha genes, and associates with GOA-1 and GPR-1. Together, this suggests that LIN-5, GPR and RIC-8 protein complexes activate G protein signaling to regulate spindle functions.