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Theses Doctoral

Investigations into roles for endocytosis in LIN-12/Notch signaling and its regulation

Chan, Jessica Yu

The LIN-12/Notch signaling pathway is highly conserved in all animals, and is crucial for proper development. It is a key pathway in specifying cell fate in many cellular contexts, and dysregulation of the pathway can have deleterious consequences. Therefore, understanding how LIN-12/Notch signaling is regulated in different contexts has been a main area of interest in the field. Previous studies in different model organisms have identified many modes of regulation of the signaling pathway, one of which is endocytosis of the ligand and receptor. Here, I further investigated the role of endocytosis in LIN-12/Notch signaling in multiple developmental contexts in Caenorhabditis elegans. Work in Drosophila and vertebrates had previously established that ligand-mediated activation of Notch requires ubiquitination of the intracellular domain of the transmembrane ligand and the activity of the endocytic adaptor Epsin in the signaling cell. The consensus in the field is that Epsin-mediated endocytosis of mono-ubiquitinated ligand generates a pulling force that exposes a cleavage site in Notch for an ADAM protease, a critical step in signal transduction. In contrast, in this thesis, I examined two different transmembrane ligands in several different cell contexts and found that activation of LIN-12/Notch and the paralogous GLP-1/Notch in C. elegans does not require either Epsin-mediated endocytosis or ubiquitination of the intracellular domain of the ligand. Results obtained by a collaborator indicate that C. elegans ligand and receptor interactions are tuned to a lower force threshold than are Drosophila ligand and receptor interactions, potentially accounting for these differences.

I also looked at the role of endocytosis in regulating LIN-12 signaling in the context of vulval development. The cell fate pattern of six vulval precursor cells (VPCs) is mediated by EGFR and LIN-12/Notch signaling. Previous work using multicopy transgenes in fixed specimens indicated that LIN-12 is post-translationally downregulated via endocytosis in response to EGFR activation in the VPC named P6.p, an event that appeared essential for ligands to activate LIN-12/Notch in neighboring VPCs. In this thesis, I manipulate the endogenous lin-12 gene and examine live specimens to show that LIN-12 appears to be regulated transcriptionally in P6.p and evidence that there may be additional potential endocytic motifs that may regulate LIN-12 in this context.

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More About This Work

Academic Units
Biological Sciences
Thesis Advisors
Greenwald, Iva
Degree
Ph.D., Columbia University
Published Here
October 20, 2020