DISSECTING THE ROLE OF SUP-17/ADAM10 IN THE BONE MORPHOGENETIC PROTEIN SIGNALING PATHWAY IN C. ELEGANS

Abstract

Bone morphogenetic proteins (BMPs) belong to the transforming growth factor (TGF-b) superfamily of ligands and mediate a highly conserved signal transduction cascade. Following ligand binding, the type I receptors are phosphorylated by the type II receptors on the cell surface. Then activated type I receptors phosphorylate and release the receptor mediated Smads (R-Smads), which subsequently complex with common mediator Smad (co-Smad) and shuttle into the nucleus to regulate target gene transcription. The BMP pathway plays multiple important roles in regulating development and tissue homeostasis. Abnormalities of the BMP signaling pathway are often associated with physiological disorders and diseases in human, which include, but are not limited to, skeleton diseases, cardiovascular disorders and cancers. Thus it is important to ensure tight regulations of BMP signaling. The Liu lab uses C. elegans as a model system to investigate regulations of a BMP-like signaling pathway. Using a genetic screen highly specific to the BMP pathway, many novel modulators of the pathway have been identified that function at the ligand-receptor level. Among them are two paralogous tetraspanins, TSP-12 and TSP-14, the netrin receptor UNC- 40/Neogenin, and the RGM protein DRAG-1. However, how these new players interact with the ligand-receptor complex or with other cell surface modulators to regulate BMP signaling is not fully understood. SUP-17/ADAM10 belongs to the ADAM (a disintegrin and metalloprotease) family of transmembrane proteins that are known for cleaving many membrane and membrane-associated proteins through ‘ectodomain shedding’. In this thesis, I reveal that SUP-17/ADAM10 is expressed and functions in the signal receiving cells to regulate BMP signaling. Endogenously tagged SUP-17 is localized both to the cell surface and to intracellular vesicles. In embryos, null mutations in tsp-12, but not in tsp-14, cause decreased SUP-17 localization at the cell surface, and show increased accumulated localization in early and late endosomes. TSP-12 may also facilitate the cleavage of SUP-17’s pro-domain in gravid adults. Using genetic approaches, I identified UNC-40/Neogenin as one of the substrates of SUP-17 in BMP signaling. Using western blotting, I generated preliminary evidence suggesting that the endogenous UNC-40 may be cleaved by SUP-17, and that there is a slight increase in steady state protein level of UNC-40 in adult tsp-12(0) mutants and a decrease in drag-1(0) mutants. These results suggested a model for how these BMP modulators may function to regulate BMP signaling: tetraspanins TSP-12 and TSP-14 regulate the cell surface localization and maturation of SUP-17/ADAM10, which in turn cleaves UNC-40/neogenin and other proteins to regulate the signaling pathway. Future work will be needed to identify the additional substrates of SUP-17/ADAM10 in the BMP pathway.