Role of O-glycosylation in Animal Development

点击翻译按钮获取中文摘要

Mucin-type O-linked glycosylation is a widespread and evolutionarily conserved protein modification catalyzed by a family of enzymes (PGANTs or GalNAcTs) that transfer the sugar N-acetylgalactosamine (GalNAc) to the hydroxyl group of serines and threonines in proteins that are destined to be membrane-bound or secreted. Defects in this type of glycosylation are responsible for the human diseases familial tumoral calcinosis and Tn syndrome. Additionally, changes in O-glycosylation have been associated with tumor progression and metastasis. More recently, genome-wide association studies have identified the genes encoding the enzymes that are responsible for initiating O-glycosylation among those associated with HDL-cholesterol levels, triglyceride levels, congenital heart defects, colon cancer and bone mineral density. From these studies, it is apparent that this conserved protein modification has a multitude of biological roles. The focus of our research group is to elucidate the mechanistic role of O-glycans during development in order to understand how they contribute to disease susceptibility and progression. Previous work from our group demonstrated that O-linked glycosylation is essential for viability in Drosophila. Moreover, we have demonstrated that the enzymes responsible for the initiation of O-glycosylation are required in specific tissues, suggesting unique tissue-specific functions. For example, we demonstrated that one essential PGANT is responsible for proper gut function. Loss of this glycosyltransferase resulted in reduced secretion of O-glycosylated proteins into the lumen of the gut and morphological alterations in the cells responsible for gut acidification. Ongoing work centers on defining the mechanistic roles for O-glycans in specific cells within the digestive system. We have evidence for crucial roles for O-glycosylation in the proper function of diverse cell types within the digestive tract. These studies have implications for the role of this protein modification in the proper gut function in higher eukaryotes, as these genes are abundantly expressed in the stomach, small intestine and colon of mice and humans. In other work, we found that loss of one PGANT family member alters secretion of an ECM protein, thereby influencing BM composition and disrupting integrin-mediated cell adhesion during Drosophila wing development. Likewise, we demonstrated that O-glycosylation also modulates the composition of the ECM during mammalian organ development, influencing integrin and FGF signaling, thereby affecting cell proliferation and growth of the developing salivary glands. These results highlight a conserved role for O-glycosylation in the establishment of cellular microenvironments and have implications for the role of this protein modification in both development and disease. We are currently collaborating with the Tabak laboratory to further investigate the effects of loss of O-glycosylation on other aspects of mammalian development. Specifically, we have found that heart development is compromised in Galnt1-deficient mice. We are currently defining the cellular and molecular changes occurring in the Galnt1-deficient hearts and identifying the direct targets of Galnt that may be responsible for mediating these effects. Finally, we have developed a system for real-time imaging of O-glycosylation and polarized secretion in a living organ. Our previous studies have demonstrated that the loss of certain O-glycosyltransferases affects secretion in Drosophila cells and alters Golgi organization, suggesting a role for these enzymes in maintaining proper secretory apparatus structure. As O-glycosyltransferases are components of the secretory apparatus and responsible for the modification of secreted and membrane bound proteins, we hypothesize that they play crucial roles in conserved biological functions related to secretion, transport, stability and/or function of proteins. We are investigating how the PGANTs are mediating effects on secretion and secretory apparatus structure by imaging specific secretory events in real time in organs where certain family members have been mutated. Thus far, we have evidence for the role of a number of family members in specific aspects of polarized secretion. In summary, we are using information gleaned from Drosophila to better focus on crucial aspects of development affected by O-glycosylation in more complex mammalian systems. We are also using real-time imaging within living organs to define the specific processes by which O-glycosylation influences secretion. Our hope is that the cumulative results of the studies described above will elucidate the mechanisms by which this conserved protein modification operates in both normal development and in disease susceptibility.