title: Cloning of a mouse beta 1,3 N-acetylglucosaminyltransferase GlcNAc(beta 1,3)Gal(beta 1,4)Glc-ceramide synthase gene encoding the key regulator of lacto-series glycolipid biosynthesis.
abstract: The distinction between the different classes of glycolipids is conditioned by the action of specific core transferases. The entry point for lacto-series glycolipids is catalyzed by the beta1,3 N-acetylglucosaminyltransferase GlcNAc(beta1,3)Gal(beta1,4)Glc-ceramide (Lc3) synthase enzyme. The Lc3 synthase activity has been shown to be regulated during development, especially during brain morphogenesis. Here, we report the molecular cloning of a mouse gene encoding an Lc3 synthase enzyme. The mouse cDNA included an open reading frame of 1131 base pairs encoding a protein of 376 amino acids. The Lc3 synthase protein shared several structural motifs previously identified in the members of the beta1,3 glycosyltransferase superfamily. The Lc3 synthase enzyme efficiently utilized the lactosyl ceramide glycolipid acceptor. The identity of the reaction products of Lc3 synthase-transfected CHOP2/1 cells was confirmed by thin-layer chromatography immunostaining using antibodies TE-8 and 1B2 that recognize Lc3 and Gal(beta1,4)GlcNAc(beta1,3)Gal(beta1,4)Glc-ceramide (nLc4) structures, respectively. In addition to the initiating activity for lacto-chain synthesis, the Lc3 synthase could extend the terminal N-acetyllactosamine unit of nLc4 and also had a broad specificity for gangliosides GA1, GM1, and GD1b to generate neolacto-ganglio hybrid structures. The mouse Lc3 synthase gene was mainly expressed during embryonic development. In situ hybridization analysis revealed that that the Lc3 synthase was expressed in most tissues at embryonic day 11 with elevated expression in the developing central nervous system. Postnatally, the expression was restricted to splenic B-cells, the placenta, and cerebellar Purkinje cells where it colocalized with HNK-1 reactivity. These data support a key role for the Lc3 synthase in regulating neolacto-series glycolipid synthesis during embryonic development.
authors.affiliation: Eunice Kennedy Shriver Center, University of Massachusetts Medical School, Waltham, Massachusetts 02452, USA.
authors.forename: T R
authors.initials: TR
authors.lastname: Henion
authors.forename: D
authors.initials: D
authors.lastname: Zhou
authors.forename: D P
authors.initials: DP
authors.lastname: Wolfer
authors.forename: F B
authors.initials: FB
authors.lastname: Jungalwala
authors.forename: T
authors.initials: T
authors.lastname: Hennet
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000606:Amino Sugars / Q000737:chemistry; D000818:Animals; D001402:B-Lymphocytes / Q000378:metabolism; D001483:Base Sequence; D015152:Blotting, Northern; D001921:Brain / Q000196:embryology / Q000378:metabolism; D002460:Cell Line; D002531:Cerebellum / Q000378:metabolism; D002855:Chromatography, Thin Layer; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D006017:Glycolipids / Q000096:biosynthesis*; D017403:In Situ Hybridization; D007313:Insecta; D051379:Mice; D008956:Models, Chemical; D008969:Molecular Sequence Data; D017351:N-Acetylglucosaminyltransferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D016366:Open Reading Frames; D011994:Recombinant Proteins / Q000378:metabolism; D013154:Spleen / Q000166:cytology; D013379:Substrate Specificity; D013997:Time Factors; D014018:Tissue Distribution
chemical_list: D000606:Amino Sugars; D018076:DNA, Complementary; D006017:Glycolipids; D011994:Recombinant Proteins; C000458:N-acetyllactosamine; D017351:N-Acetylglucosaminyltransferases; C067641:lactosylceramide 1,3-N-acetyl-beta-D-glucosaminyltransferase