[RNW] - Gamete Interactions in Caenorhabditis elegans

RELEVANCE: Fertilization is a biological process with important medical, social and economic implications. From extensive study, the events of fertilization are known in some detail. However, the molecular underpinnings of these events generally remain elusive. Most previous work on fertilization has relied on biochemical and immunological approaches. Our work is groundbreaking in the application of classic gentic analysis to this vital area of research. RATIONALE: Many of the genetic and molecular tools developed for C. elegans are not available or are very difficult to utilize in other organisms traditionally used for studying fertilization. One of the most significant advantages of C. elegans is the ability to isolate and maintain mutants that affect sperm or eggs and no other cells. Previously, through the study of sterile mutants, we have identified some of the first molecules required for productive gamete interactions in C. elegans. We propose to gain a better understanding how the spe-13 and egg-7 genes function to ensure successful fertilization. Mutations in both these genes lead to the production of morphologically normal gametes that fail in fertilization. Futher, a full understanding of the molecular mechanisms of fertilization is impossible without a more complete inventory of molecular components. Therefore we aim to identify new fertilization molecules through genetic and molecular analysis. Particular emphasis will be placed on the identification of genes required in oocytes. OBJECTIVES: The goal of this proposal is to further our understanding of fertilization in C. elegans by conducting the following experimental aims: 1) Investigate the function, interactions, and control of the cellular distribution of SPE-13. 2) Investigate how EGG-7 impacts glycosylation and GPI-anchored protein pathways required for fertilization. We will characterize human pathogenic variants for their effect on C. elegans EGG-7 function. 3) Conduct a CRISPR/Cas9 engineered balancer-based genetic screen for new fertilization mutants and determine the molecular nature of new genes defined by a fertilization defective mutant phenotype. This work will complement fertility studies in other organisms and provide molecular insights to the diversity of reproductive strategies.