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A novel, promoter‐based, target‐specific assay identifies 2‐deoxy‐ d ‐glucose as an inhibitor of globotriaosylceramide biosynthesis
Author(s) -
Okuda Tetsuya,
Furukawa Koichi,
Nakayama Kenichi
Publication year - 2009
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/j.1742-4658.2009.07215.x
Subject(s) - globotriaosylceramide , lactosylceramide , biosynthesis , biochemistry , glycosphingolipid , glycosyltransferase , biology , microbiology and biotechnology , enzyme , chemistry , fabry disease , medicine , disease , pathology
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 α1,4‐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 by 2‐deoxy‐ d ‐glucose treatment. These results indicate that 2‐deoxy‐ d ‐glucose can control Gb3 biosynthesis through the inhibition of Gb3S transcription. Furthermore, we demonstrated the general utility of our novel screening assay for the identification of new inhibitors of glycosphingolipid biosynthesis.