Characterization of AtNST‐KT1, a novel UDP‐galactose transporter from Arabidopsis thaliana

Abstract

Nucleotide sugar transporters (NST) mediate the transfer of nucleotide sugars from the cytosol into the lumen of the endoplasmatic reticulum and the Golgi apparatus. Because the NSTs show similarities with the plastidic phosphate translocators (pPTs), these proteins were grouped into the TPT/NST superfamily. In this study, a member of the NST-KT family, AtNST-KT1, was functionally characterized by expression of the corresponding cDNA in yeast cells and subsequent transport experiments. The histidine-tagged protein was purified by affinity chromatography and reconstituted into proteoliposomes. The substrate specificity of AtNST-KT1 was determined by measuring the import of radiolabelled nucleotide mono phosphates into liposomes preloaded with various unlabelled nucleotide sugars. This approach has the advantage that only one substrate has to be used in a radioactively labelled form while all the nucleotide sugars can be provided unlabelled. It turned out that AtNST-KT1 represents a monospecific NST transporting UMP in counterexchange with UDP-Gal but did not transport other nucleotide sugars. The AtNST-KT1 gene is ubiquitously expressed in all tissues. AtNST-KT1 is localized to Golgi membranes. Thus, AtNST-KT1 is most probably involved in the synthesis of galactose-containing glyco-conjugates in plants.

The Import of S-Adenosylmethionine into the Golgi Apparatus Is Required for the Methylation of Homogalacturonan

“…The chemical structure of SAM resembles that of the nucleotide sugars (NSs), and, since different NSTs have been shown previously to be localized in the Golgi apparatus in plants (Baldwin et al, 2001;Bakker et al, 2005;Norambuena et al, 2005;Rollwitz et al, 2006), we wanted to evaluate whether the transport of SAM is also mediated by a NST-like transporter. We thus performed our SAM import assay in the presence of DIDS, a general inhibitor of the NSTs (Wulff et al, 2000).…”

Monoubiquitin-dependent endocytosis of the IRON-REGULATED TRANSPORTER 1 (IRT1) transporter controls iron uptake in plants

“…No colocalization or only very limited overlap was observed with the Golgi-localized Nucleotide Sugar Transporter (19) or with the VAM3/SYP22 prevacuolar compartment/tonoplast-localized syntaxin (20,21) (Fig. S4).…”

UUAT1 Is a Golgi-Localized UDP-Uronic Acid Transporter That Modulates the Polysaccharide Composition of Arabidopsis Seed Mucilage

“…In Arabidopsis thaliana, the genes encoding for NSTs are similar to those encoding for plastidic triose phosphate translocators (TPTs); together, 44 NSTs and 7 TPTs form a gene family of 51 members (Knappe et al, 2003;Rautengarten et al, 2014). To date, a number of these NSTs from Arabidopsis have been functionally characterized, specifically transporters for GDP-mannose (GDP-Man), GDP-fucose (GDP-Fuc), UDP-galactose (UDP-Gal), UDP-glucose (UDP-Glc), UDP-rhamnose (UDP-Rha), and UDP-xylose (UDP-Xyl) (Bakker et al, 2005;Baldwin et al, 2001;Ebert et al, 2015;Handford et al, 2012Handford et al, , 2004Norambuena et al, 2002Norambuena et al, , 2005Rautengarten et al, 2014Rautengarten et al, , 2016Rollwitz et al, 2006).…”