Identification and Characterization of a Golgi-Localized UDP-Xylose Transporter Family from Arabidopsis.

Berit Ebert,William G.T Willats,Thomas Herter,Markus Pauly,Henrik Vibe Scheller,Solomon Stonebloom,Xiaoyuan Guo,Joshua L Heazlewood,Carsten Rautengarten,Guangyan Xiong,Andreia M Smith-Moritz,Leanne Jade G Chan,Paul D Adams,Christopher J Petzold

doi:10.1105/tpc.114.133827

Abstract

Most glycosylation reactions require activated glycosyl donors in the form of nucleotide sugars to drive processes such as posttranslational modifications and polysaccharide biosynthesis. Most plant cell wall polysaccharides are biosynthesized in the Golgi apparatus from cytosolic-derived nucleotide sugars, which are actively transferred into the Golgi lumen by nucleotide sugar transporters (NSTs). An exception is UDP-xylose, which is biosynthesized in both the cytosol and the Golgi lumen by a family of UDP-xylose synthases. The NST-based transport of UDP-xylose into the Golgi lumen would appear to be redundant. However, employing a recently developed approach, we identified three UDP-xylose transporters in the Arabidopsis thaliana NST family and designated them UDP-XYLOSE TRANSPORTER1 (UXT1) to UXT3. All three transporters localize to the Golgi apparatus, and UXT1 also localizes to the endoplasmic reticulum. Mutants in UXT1 exhibit ∼30% reduction in xylose in stem cell walls. These findings support the importance of the cytosolic UDP-xylose pool and UDP-xylose transporters in cell wall biosynthesis.

Highlights

Plant cell walls are composed of various polysaccharides and with the exception of cellulose and callose, these cell wall polysaccharides are biosynthesized in the lumen of the Golgi apparatus by families of glycosyltransferases (Scheible and Pauly, 2004; Liepman et al, 2010)
nucleotide sugar transporters (NSTs) belong to the NST/triose phosphate translocator (TPT) superfamily, and the fact that they are present in all eukaryotes testifies to their biological significance (Knappe et al, 2003)
Since we observed a significant decrease in cell wall xylose in the uxt1 mutants, we focused on antibodies recognizing xylan structures, including LM10/LM11, UX1, and AX1 [which was produced against arabinose-substituted b-(1,4)xylan from wheat (Triticum aestivum)]

Summary

INTRODUCTION

Plant cell walls are composed of various polysaccharides and with the exception of cellulose and callose, these cell wall polysaccharides are biosynthesized in the lumen of the Golgi apparatus by families of glycosyltransferases (Scheible and Pauly, 2004; Liepman et al, 2010). To assess if mutations in the UXTs affect the biosynthesis of specific polysaccharides, we prepared alcohol insoluble residue (AIR) from leaves, flowers, and young (upper) and mature (lower) inflorescence stem tissue from 6- to 8-week-old plants and analyzed the monosaccharide composition (Supplemental Table 2). The Xyl content in mature inflorescence stems from the uxt mutants was decreased by 16% (Figure 5B), whereas a reduction of 34% was observed in young stem tissue (Figure 5A) These data confirm the importance of UXT1 for the biosynthesis of Xyl-containing cell wall polymers. A significant reduction in cell wall GlcA content of ;25 to 37% was observed in mature and young parts of the inflorescence stems from uxt mutants. Our results indicate that the UXTs do not have an obvious effect on protein N-glycosylation

DISCUSSION

Conclusions

METHODS

Findings

Cloning Procedures