STUDY ON THE BIOLOBICAL AND PHYSIOLOGICAL FUNCTION OF 6-O-SULFATION IN HEPARANSULFATE/HEPARIN

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Heparan sulfate (HS) plays critical roles in a variety of developmental, physiological, and pathogenic processes. These functions are due to interact in a structure-dependent manner with numerous growth factors that participate in cellular signaling. We examined the roles of 2-O-sulfate and 6-O-sulfate in HS using mice, chick, drosophila, and mutant cells.1.Regulation of heparin-binding growth factor (HB-GF)-induced signaling by heparan sulfate(1) We indicated that VEGF165-dependent tube formation and mitogenic activity was enhanced by 6-O-sulfate in heparan sulfate (2005, J.Biol.Chem.). (2) HS6ST-1 knockout mice exhibited the abnormal angiogenesis in placental labyrinthine zone. These defects appear to be due to the reduced expression of VEGF (2007, J.Biol.Chem.) (3) In drosophila, FGF signaling regulates tracheal system formation. 39% of Hs6st null embryos exhibited tracheal defects (2006, J.Cell.Biol.). (4) The formation of limb bud involves various HB-GFs. When HS2ST in the prospec … More tive limb of chick embryo was inhibited by siRNA, the limb buds were truncated and the expression of FGF-8 was reduced in the apical ectodermal ridge (2007, J.Biol.Chem.). (5) We examined the functions of HS 6-O-sulfation in HB-GFs signaling using fibroblast derived from HS6ST-1 ; HS6ST-2 double mutant mice (dKO-MEF). In dKO-MEFs, FGF-4-and FGF-2-dependent signaling was reduced to approximately 30% and 60% of WT-MEFs, respectively. FGF-1-dependent signaling was moderately reduced compared to that of WT-MEFs but only at lower FGF-1 concentration. The results suggest that 6-O-sulfate in HS probably regulates the signaling of some of HB-GFs, including FGFs, by inducing the differential interaction between ligands and their receptors (in submission).2.Biosynthesis of 6-O-sulfation in heparinMast cells have many proteases, which are bound to heparin in granules, and released by the interaction of the antigen with cell surface IgE. Sulfaion of Heparin is required for the interaction with some of these proteins in granules. Therefore, it is important to elucidate biosynthesis pathway of 6-O-sulfation of heparin. Structural analysis of heparin derived from ears of knockout mice showed that HS6ST1 null mice generated normal sulfated heparin. On the other hand, heparin derived from HS6ST-2 null mice was reduced to 50% of 6-O-sulfation in normal heparin. We are investigating the structure changes of heparin produced by the double knockout mice using cultured mast cell from embryonic skin. Less