tailieunhanh - Báo cáo khoa học: A novel, promoter-based, target-specific assay identifies 2-deoxy-D-glucose as an inhibitor of globotriaosylceramide biosynthesis

Abnormal biosynthesis of globotriaosylceramide (Gb3) is known to be associated with Gb3-related diseases, such as Fabry disease. The Gb3 synthase gene (Gb3S) codes for a1,4-galactosyltransferase, which is a key enzyme involved in Gb3 biosynthesisin vivo. | A novel promoter-based target-specific assay identifies 2-deoxy-D-glucose as an inhibitor of globotriaosylceramide biosynthesis Tetsuya Okuda1 Koichi Furukawa2 and Ken-ichi Nakayama1 1 Glycolipids Function Analysis Team Health Technology Research Center National institute of Advanced Industrial science and Technology Kagawa Japan 2 Department of Biochemistry II Graduate Schoolof Medicine Nagoya University Aichi Japan Keywords Fabry disease glycosphingolipid glycosyltransferase hemolytic uremic syndrome promoter Correspondence T. Okuda Glycolipids Function Analysis Team Health Technology Research Center National institute of Advanced Industrial Science and Technology AIST 2217-14 Hayashi Takamatsu Kagawa 761-0395 Japan Fax 81 87 869 3593 Tel 81 87 869 3563 E-mail t-okuda@ Received 27 April2009 revised 22 June 2009 accepted 15 July 2009 doi Abnormal biosynthesis of globotriaosylceramide Gb3 is known to be associated with Gb3-related diseases such as Fabry disease. The Gb3 synthase gene Gb3S codes for a1 4-galactosyltransferase which is a key enzyme involved in Gb3 biosynthesis in vivo. Transcriptional repression of Gb3S is a way to control Gb3 biosynthesis and may be a suitable target for the treatment of Gb3-related diseases. To find a transcriptional inhibitor for Gb3S we developed a convenient cell-based chemical screening assay system by constructing a fusion gene construct of the human Gb3S promoter and a secreted luciferase as reporter. Using this assay we identified 2-deoxy-D-glucose as a potent inhibitor for the Gb3S promoter. In cultured cells 2-deoxy-D-glucose markedly reduced endogenous Gb3S mRNA levels resulting in a reduction in cellular Gb3 content and a corresponding accumulation of the precursor lactosylceramide. Moreover cytokine-induced expression of Gb3 on the cell surface of endothelial cells which is closely related to the onset of hemolytic uremic syndrome in O157-infected patients was also suppressed .