UBC Theses and Dissertations
Characterization of starfish yolk and cortical granule proteins, and of a novel extracellular proteoglycan implicated in digestive tract morphogenesis Reimer, Corinne L.
Extracellular matrix (ECM) is thought to play a major role in morphogenesis by influencing processes such as cell migration and differentiation, although the specific mechanisms involved are poorly understood. This study examined the ECM and egg storage granules of starfish (Pisaster ochraceus) embryos, and attempted to identity components important for digestive tract morphogenesis. Three monoclonal antibodies were developed with specificities for the ECM, yolk and cortical granules in Pisaster eggs and embryos. These antibodies were then used to localize, isolate and characterize the antigens through early development, using immunohistochemistry, immunocytochemistry, immunochemical and biochemical techniques. The first antibody, PM1, binds to a large extracellular proteoglycan, which appears in the blastocoel matrix at mid-gastrulation, and is synthesized only by endodermally-derived tissues. The use of PM1 antibody as a function blocking agent in live embryo cultures suggested that it plays an important role in digestive tract morphogenesis. A second antibody recognizes a protein localized in cortical granules of unfertilized eggs. The majority of these granules are located in the peripheral egg cytoplasm and are released at fertilization. However, a second morphologically identical population of granules remain dispersed throughout the egg cytoplasm, and appear to contribute to ECMs of the developing embryo, including the blastocoel ECM, basement membranes, and the hyaline layer. The function of this protein is currently unknown; however, it has a different storage and secretion profile from the PM1 proteoglycan, suggesting its role in the blastocoel matrix may be different. A third antibody recognizes proteins stored in yolk granules located throughout the egg and cells of the developing embryo, which do not appear to contribute to ECMs during embryogenesis. Partial biochemical characterizations using the anti-yolk antibody revealed that there are several molecular species of yolk proteins present in the oocyte, and that their molecular composition changes during embryogenesis. Depletion of the yolk proteins is not significant until the larval stage, suggesting that they do not play a major role until later in development.
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