Biological function of keratan sulfate glycosaminoglycan for corneal extracellula

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Keratan sulfate (KS) proteoglycan (PG) is one of the major components of the cornea, and suggested to have important roles for corneal development and maintenance of transparency of the tissue. KS-PG is consists of two different components, core proteins and attached carbohydrates called glycosaminoglycans (GAGs). It has been postulated that KS-PG core proteins bind to collagen fibrils and regulate diameter of the fibrils, and KS-GAGs regulate interfibrillar spacing. By using several approaches including gene-knockout mice, importance of KS-PG core proteins, such as lumican and keratocan, over corneal development has been well investigated, however, biological function of KS-GAG chains has not been extensively studied, since biosynthetic pathway of the carbohydrate has not been established. KS-GAG is a linearly elongated carbohydrate chain that consists of repeating disaccharide units of 3Gal~1-4GlcNAc~1- (Gal, galactose; GlcNAc, Nacetylglucosamine), with sulfate on 6-0 position of Gal and GlcNAc. We have studied biosynthesis of corneal KS carbohydrate, and identified that 4 Golgi-Ioca/ized enzymes, ~1 ,3-N-acetylglucosaminyltransferase 7, ~1 ,4-galactosyltransferase 4, KS galactose 6-0 sulfotransferase and corneal N-acetylglucosamine 6-0 sulfotransferase, are required for KS-GAG production in vitro as well as in vivo. We also found that lack of sulfation of KS carbohydrate alters corneal extracellular matrix (ECM) structure in mouse cornea, and demonstrated functional importance of KS-GAG chain in the cornea. Based on these findings, we hypothesized that molecular structure, such as length and sulfation degree, of KS carbohydrate affects to the formation of organized corneal ECM structure. In other words, we may be able to remodel the corneal ECM structure by manipulating KS carbohydrate structure, which can be controlled by suppression and/or induction of enzymatic activity required for KS synthesis in corneal cells. To test this hypothesis, we will work on the two aims.