tailieunhanh - Báo cáo Y học: Regulation of a1,3galactosyltransferase expression in pig endothelial cells Implications for xenotransplantation

The disaccharide galactosea1,3galactose (theaGal epitope) is the major xenoantigen responsible for the hyperacute vascular rejection occurring in pig-to-primates organ trans-plantation. The synthesis of theaGal epitope is catalyzed by the enzyme a1,3-galactosyltransferase (a1,3GalT). To be able to control porcinea1,3GalT gene expression speci®c-ally, we have analyzed the upstreamportionof thea1,3GalT gene, and identi®ed the regulatory sequences. | Eur. J. Biochem. 269 1464-1473 2002 FEBS 2002 Regulation of a1 3galactosyltransferase expression in pig endothelial cells Implications for xenotransplantation Dominiaue Mercier1 2 Beatrice Charreau3 Anne Wierinckx2 Remco Keiiser1. Lize Adriaensens1 Bfl w BB B B B B B B B B BB B B B B BB BB w B B B B BB w B BB B B B B B WBB B B BB B B B BB BB B w BB B B B BB B B B B B BB B B BB BB B B B BB B B B Renate van den Berg1 and David H. Joziasse1 1 Department of Molecular Cell Biology Research Institute of Immunology and Inflammatory Diseases and department of Medical Pharmacology Research Institute Neurosciences VU VUmc Amsterdam the Netherlands 3Institut de Transplantation et de Recherche en Transplantation ITERT INSERM U437 Nantes France The disaccharide galactosea1 3galactose the aGal epitope is the major xenoantigen responsible for the hyperacute vascular rejection occurring in pig-to-primates organ transplantation. The synthesis of the aGal epitope is catalyzed by the enzyme a1 3-galactosyltransferase a1 3GalT . To be able to control porcine a1 3GalT gene expression specially we have analyzed the upstream portion of the a1 3GalT gene and identified the regulatory sequences. Porcine a1 3GalT transcripts were detected by 5 RACE analysis and the corresponding genomic sequences were isolated from a phage library. The porcine a1 3GalT gene consists of at least 10 different exons four of which contain 5 untranslated sequence. Four distinct promoters termed AD drive a1 3GalT gene transcription in porcine cells. A combination of alternative promoter usage and alternative splicing produces a series of transcripts that differ in their 5 portion but encode the same protein. Promoters A-C have been isolated and functionally characterized using luciferase reporter gene assays in transfected porcine endothelial cells PEC-A . Promoter preference in porcine endothelial cells was estimated on the basis of relative transcript levels as determined by real-time quantitative PCR. More .