Paper- Restricted to Campus Access
Michelle Vande Berg
Caenorhabditis elegans are microscopic worms that are a great model organism for examining genes and proteins involved in the reproductive system because of their fecundity, easy manipulation, and visible gonad. The Lyczak lab has studied two versions of a puromycin aminopeptidase known as PAM-1. We have identified a missense mutation pam-1(or347), a partial loss-of-function of pam-1 and a nonsense mutation, pam-1(or403), a complete loss of function. This PAM-1 protein is 36 – 37% similar to the humans Npepps protein. pam-1 mutants lay fewer viable embryos than wild-type worms because they have trouble completing meiosis. However, we have also found another gene that rescues pam-1 mutants' low hatch rate, known as wee-1.3. WEE-1.3 is involved in oocyte maturation. It inhibits the maturation-promoting factor (MPF) in oocytes to keep the oocytes immature prior to fertilization. A suppressing mutation in wee-1.3 raised the hatch rate of pam-1 embryos. Due to this, we believe PAM-1 has a direct or indirect effect on MPF. MPF is made up of cyclin B and cyclin-dependent kinase 1(CDK-1). To examine this interaction, I have conducted localization studies of CDK-1 in wild type, pam-1(or347), and pam-1(or403) worms using the confocal. I have also performed interaction studies of PAM-1 with MPF components in oocytes by using CDK-1 and WEE-1.3 RNAi with different variations of the strains. With these in hand, we have further characterized PAM-1 and MPF’s interaction. RNAi experiments have shown that pam-1(or347) has protection from the knockout of CDK-1. The localization studies have shown a significant difference between wild type’s and pam-1 mutants CDK-1 intensity, confirming that more CDK-1 is found at higher levels in the pam-1(or347) worms. To examine if CDK-1’s intensity changes based upon WEE–1.3 activation state, WEE-1.3 RNAi experiments were done upon worms with CDK-1 GFP. However, there was not much of significant difference found. Overall, pam-1(or347) mutants has shown that it has protection from WEE-1.3 knockout and protection from the loss of CDK-1 as well as a role in regulating CDK-1 levels. However, WEE-1.3 does not localize CDK-1. While the data supports a role for PAM-1 in regulating the MPF, future studies will be needed to determine how missense and nonsense mutations differentially affect the results.
Davis, Quaran, "Genetic Interactions of PAM-1 & MPF in Developing Oocytes" (2022). Biology Honors Papers. 50.