The role of PAM-1 in the regulation of the cell cycle and polarity establishment in C. elegans

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PROJECT SUMMARY The cell cycle is a tightly regulated process controlled by the transient activation of cyclin- dependent kinases (CDKs). Activation and deactivation of CDKs is coordinated by its associated cyclin as well as other kinases, phosphatases, and the degradation machinery. Coordination of these complexes drives the cell through each stage of the cell cycle and is necessary for development, tissue maintenance, and fertility. Puromycin-sensitive aminopeptidases (PSAs) are highly conserved metalloproteases implicated in cell-cycle regulation in numerous organisms. Despite widespread roles in this process, the mechanism by which PSAs interact with the cyclin/CDKs and other cell-cycle machinery is poorly understood. A key goal of our project is to uncover these interactions in the C. elegans model. In our previous work, we showed that the C. elegans PSA homolog, PAM-1, plays a role in meiotic exit regulation and anterior-posterior axis establishment and when mutated, results in embryonic lethality. In a screen for suppressors of pam-1, we identified a mutation in wee-1.3 that rescues the lethality of pam-1 mutants. WEE-1.3 is an inhibitory kinase that negatively regulates CDK-1, part of the maturation promoting complex. We went on to show that pam-1 and wee-1.3 genetically interact in polarity establishment and oocyte maturation, suggesting a broad role for PAM-1 in regulating the cell cycle. The proposed work seeks to further characterize the role of PAM-1 and its interaction with WEE-1.3 and the MPF, testing the hypothesis that PAM-1 is necessary for full MPF activity. Work will focus on characterization of oocyte maturation, meiosis, and mitosis in the early embryo through a combination of time-lapse imaging, genetic interaction, and localization studies. In addition, we will characterize and identify the genes mutated in additional suppressors of pam-1 to find new interactions. We expect to gain a new understanding of how PAM-1 regulates that cell cycle that is likely to be applicable to other systems. The proposed experiments will be largely carried out by undergraduates who will be mentored to provide them the guidance and expertise needed for success in science careers and graduate work. Inclusion and mentoring of students underrepresented in STEM is an important component of this work.