Submission Date

4-26-2021

Document Type

Paper- Restricted to Campus Access

Department

Biology

Adviser

Rebecca Lyczak

Committee Member

Kathryn Goddard

Committee Member

Lisa Grossbauer

Department Chair

Rebecca Lyczak

Project Description

To develop into mature organisms, fertilized embryos must go through phases of growth, DNA replication, and cell division. The phases together make up the cell cycle, and movement through these phases is facilitated by many proteins. In the model organism Caenorhabditis elegans, one of these proteins is PAM-1, a cytosolic puromycin-sensitive aminopeptidase that regulates proteins necessary for the cell to progress out of meiosis. Interestingly, there is evidence that PAM-1 interacts with a protein called WEE-1.3, as the absence of PAM-1 is lethal to embryos, but a simultaneous mutation affecting WEE-1.3 rescues the embryos and somewhat increases the hatch rate. WEE-1.3 is a cell cycle regulator and prevents the premature activation of the Maturation Promoting Factor (MPF) in immature oocytes. The MPF is a complex of two proteins: CDK-1 and CYB-3. The mechanism of the interaction between PAM-1 and other proteins has yet to be fully understood. This study considers the involvement of the MPF with PAM-1 and WEE-1.3. To test if PAM-1 and the MPF interact during oocyte maturation or meiosis, we inactivated cdk-1 or cyb-3 in wild-type and pam-1 mutants. We observed significant differences between wildtype and pam-1 oocyte maturation after RNAi treatments that knocked down MPF expression, suggesting that the mutant strain pam-1(or403) may protect against loss of CDK-1 and CYB-3, with an even stronger protection by another mutant strain, pam-1(or347). We observed that there was a similar protection against the pam-1 defect of one cell arrest when CDK-1 was inactivated in both mutant strains. We also chose to observe CDK-1 localization in both pam-1 and wildtype strains, which was made possible by the creation of strains that included both GFP tagged CDK-1 and a pam-1 mutation. These strains were created and are pending verification. At first observation, CDK-1 localization seems to be severely impacted in the CDK-1::GFP strain with the pam-1(or403) mutation but not in the CDK-1::GFP strain with the pam-1(or347) mutation. Our work suggests that PAM-1 acts downstream of the MPF and WEE-1.3 kinase in regulation of oocyte maturation and meiosis, as absence of PAM-1 makes up for deficiencies in MPF components.

Comments

Supported by a National Institutes of Health (NIH) grant: GM110614

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