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Glycobiology Advance Access published online on September 29, 2004
Glycobiology, doi:10.1093/glycob/cwh159
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Received July 16, 2004
Revised September 21, 2004
Accepted September 23, 2004

ORIGINAL ARTICLES

Molecular cloning of two Arabidopsis UDP-galactose transporters by complementation of a deficient chinese hamster ovary cell line

Hans Bakker 1*, Françoise Routier 2, Stefan Oelmann 2, Wilco Jordi 3, Arjen Lommen 4, Rita Gerardy-Schahn 2, and Dirk Bosch 3

1 Plant Research International, Wageningen University and Research Centre, P.O.Box 16, 6700AA, Wageningen, The Netherlands; Zelluläre Chemie, Medizinische Hochschule Hannover, Carl-Neuberg-Strasse 1, 30625 Hannover, Germany
2 Zelluläre Chemie, Medizinische Hochschule Hannover, Carl-Neuberg-Strasse 1, 30625 Hannover, Germany
3 Plant Research International, Wageningen University and Research Centre, P.O.Box 16, 6700AA, Wageningen, The Netherlands
4 Institute of Food Safety (RIKILT), Wageningen University and Research Centre, P.O. Box 230, 6700AE Wageningen, The Netherlands

* To whom correspondence should be addressed. E-mail: Bakker.Hans{at}mh-hannover.de.


  Abstract

Nucleotide-sugar transporters (NSTs) form a family of structurally related transmembrane proteins that transport nucleotide-sugars from the cytoplasm to the ER and Golgi lumen. In these organelles, activated sugars are substrates for various glycosyltransferases involved in oligo- and polysaccharide biosynthesis. The Arabidopsis thaliana genome contains more than 40 members of this transporter gene family, of which only a few are functionally characterised. In this study, two Arabidopsis UDP-galactose transporter cDNAs (UDP-GalT1 and UDP-GalT2) are isolated by expression cloning, using a Chinese hamster ovary cell line (CHO-Lec8) that is deficient in UDP-galactose transport. The isolated genes show only 21% identity to each other and very limited sequence identity with human and yeast UDP-galactose transporters and other NSTs. Despite this low overall identity, the two proteins clearly belong to the same gene family. Besides complementing Lec8 cells, the two NSTs are shown to transport exclusively UDP-galactose by an in-vitro nucleotide-sugar transport assay. The most homologous proteins with known function are plant transporters that locate in the inner chloroplast membrane and transport triose-phosphate, phosphoenolpyruvate, glucose-6-phosphate and xylulose 5-phosphate. Also the latter proteins are members of the same family, which therefore has been named the Nucleotide-Sugar Transporter/Triose-Phosphate Transporter (NST-TPT) family.

Keywords: Cell Wall; Expression cloning; Golgi; Membrane; Nucleotide-Sugar-Transporter.
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