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Molecular Cloning of a Rat Brain cDNA, with Homology to a Tyrosine Kinase Substrate, that Induces Galactosylceramide Expression in COS‐7 Cells
Author(s) -
Ogura Kiyoshi,
Kohno Kenji,
Tai Tadashi
Publication year - 1998
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.1998.71051827.x
Subject(s) - complementary dna , microbiology and biotechnology , biology , transfection , tyrosine kinase , biochemistry , gene , signal transduction
A rat brain cDNA clone has been isolated, using a eukaryotic cell transient expression system in conjunction with an anti‐galactosylceramide (anti‐GalCer) monoclonal antibody that induces GalCer expression in COS‐7 cells. The protein was designated as GalCer expression factor‐1 (GEF‐1). A good correlation between GalCer expression and the level of the enzyme activity of UDP‐galactose:ceramide galactosyltransferase (CGT) was demonstrated. The cDNA insert encoded a polypeptide of 771 amino acids with a calculated molecular mass of 85,787 Da. The cDNA hybridized to a single mRNA of 3.1 kb in all rat organs examined, including brain, testis, and skeletal muscle. The cDNA product was determined to be a tyrosine‐phosphorylated protein with a molecular mass of 110 kDa in transfected COS‐7 cells and adult rat brain. COS‐7 cells transfected with the cDNA clone showed dramatic morphological changes: The transfected cells appeared to be fibroblast‐like cells, whereas the parent COS‐7 cells were typical epithelial‐like cells. The deduced amino acid sequences revealed a strikingly high homology to a mouse hepatocyte growth factor‐regulated tyrosine kinase substrate but no homology to CGT. Taking these results together, it is suggested that GEF‐1 may play an important role in regulating GalCer expression in the brain.