Plant Cell Advance Online Publication
Published on July 13, 2007; 10.1105/tpc.107.052985
OPEN ACCESS ARTICLE
Received May 21, 2007
Returned for revision June 22, 2007
Accepted June 25, 2007
A Unique  1,3-Galactosyltransferase Is Indispensable for the Biosynthesis of N-Glycans Containing Lewis a Structures in Arabidopsis thaliana
Richard Strasser 1*, Jayakumar Singh Bondili 2, Ulrike Vavra 1, Jennifer Schoberer 1, Barbara Svoboda 1, Josef Glössl 1, Renaud Léonard 3, Johannes Stadlmann 3, Friedrich Altmann 3, Herta Steinkellner 1, and Lukas Mach 1
1 Institute of Applied Genetics and Cell Biology, BOKU, University of Natural Resources and Applied Life Sciences, A-1190 Vienna, Austria
2 Institute of Applied Genetics and Cell Biology, BOKU, University of Natural Resources and Applied Life Sciences, A-1190 Vienna, Austria; Department of Chemistry, BOKU, University of Natural Resources and Applied Life Sciences, A-1190 Vienna, Austria
3 Department of Chemistry, BOKU, University of Natural Resources and Applied Life Sciences, A-1190 Vienna, Austria
* To whom correspondence should be addressed. E-mail: richard.strasser{at}boku.ac.at.
In plants, the only known outer-chain elongation of complex N-glycans is the formation of Lewis a [Fuc 1-4(Gal 1-3)GlcNAc-R] structures. This process involves the sequential attachment of 1,3-galactose and 1,4-fucose residues by 1,3-galactosyltransferase and 1,4-fucosyltransferase. However, the exact mechanism underlying the formation of Lewis a epitopes in plants is poorly understood, largely because one of the involved enzymes, 1,3-galactosyltransferase, has not yet been identified and characterized. Here, we report the identification of an Arabidopsis thaliana 1,3-galactosyltransferase involved in the biosynthesis of the Lewis a epitope using an expression cloning strategy. Overexpression of various candidates led to the identification of a single gene (named GALACTOSYLTRANSFERASE1 [GALT1]) that increased the originally very low Lewis a epitope levels in planta. Recombinant GALT1 protein produced in insect cells was capable of transferring 1,3-linked galactose residues to various N-glycan acceptor substrates, and subsequent treatment of the reaction products with 1,4-fucosyltransferase resulted in the generation of Lewis a structures. Furthermore, transgenic Arabidopsis plants lacking a functional GALT1 mRNA did not show any detectable amounts of Lewis a epitopes on endogenous glycoproteins. Taken together, our results demonstrate that GALT1 is both sufficient and essential for the addition of 1,3-linked galactose residues to N-glycans and thus is required for the biosynthesis of Lewis a structures in Arabidopsis. Moreover, cell biological characterization of a transiently expressed GALT1-fluorescent protein fusion using confocal laser scanning microscopy revealed the exclusive location of GALT1 within the Golgi apparatus, which is in good agreement with the proposed physiological action of the enzyme.
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