Analysis of genetic enhancers of Ras/MAPK signaling in C. elegans

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Development of a multicellular animal from a single cell is a highly complex process requiring cell coordination and communication. Several highly-conserved signal transduction pathways are repeatedly used to regulate many aspects of development. In the nematode C. elegans, Ras/MAPK signaling regulates several developmental events including meiotic progression of germ cells, oocyte growth, cell fate specification and cell migration. For Ras/MAPK signaling to elicit different biological events, there must be unique regulators and downstream targets that tailor signaling such that the appropriate outcomes are reached. We identified several positive regulators of Ras/MAPK signaling during the specification of the excretory duct cell (EDC). The EDC is one of three tubular cells that make up the excretory system. Animals missing the EDC accumulate waste fluid and die as larvae. Most of these positive regulators promote Ras/MAPK signaling specifically in the EDC. However, UBC-25 appears to have a second role in promoting oocyte growth with Ras/MAPK signaling. The objective of our NSERC research program is to understand the mechanisms by which these cell type specific regulators promote Ras/MAPK signaling. The objectives of this proposal are 1) characterize the role of UBC-25 and Ras/MAPK signaling in oocyte growth, 2) characterize the role of EKL-7 in specification of the EDC and 3) determine the molecular identities of uncloned regulators. UBC-25 is a conserved E2 ubiquitin-conjugating enzyme. In addition to specifying the EDC fate, UBC-25 functions with Ras/MAPK signaling to regulate oocyte growth, but not meiotic progression providing a unique opportunity to determine the mechanisms by which Ras/MAPK promotes oocyte growth. We hypothesize that UBC-25 antagonizes a negative regulator of Ras/MAPK signaling to regulate oocyte growth and the EDC fate. EKL-7 lacks known functional domains, but has a cluster of MAPK Erk phosphorylation sites. EKL-7 genetically interacts with known Erk substrates. We hypothesize that EKL-7 is a direct target of Ras/MAPK signaling that cooperates with other Erk substrates to specify the EDC fate. Finally, we will complete genetic mapping of two novel mutations in genes that promote Ras/MAPK signaling in the EDC to determine their molecular identities to determine how they provide specificity to signaling. This research will provide new insights into the molecular functions of UBC-25 and EKL-7 proteins, the mechanisms that regulate Ras/MAPK signaling, and how Ras/MAPK regulates oocyte growth. This work will train HQP in critical thinking, model building, scientific writing, and molecular, genetic, and cell biological skills and knowledge for careers in the biological sciences and engineering.