Abstract
GDP-d-mannose (GDP-d-Man) is an important intermediate in ascorbic acid (AsA) synthesis, cell wall synthesis, protein N-glycosylation, and glycosylphosphatidylinositol-anchoring in plants. Thus, the modulation of intracellular levels of GDP-d-Man could be important for maintaining various cellular processes. Here an Arabidopsis GDP-d-Man pyrophosphohydrolase, AtNUDX9 (AtNUDT9; At3g46200), which hydrolysed GDP-d-Man to GMP and mannose 1-phosphate, was identified. The K m and V max values for GDP-d-Man of AtNUDX9 were 376±24 μM and 1.61±0.15 μmol min(-1) mg(-1) protein, respectively. Among various tissues, the expression levels of AtNUDX9 and the total activity of GDP-d-Man pyrophosphohydrolase were the highest in the roots. The GDP-d-Man pyrophosphohydrolase activity was increased in the root of plants grown in the presence of ammonium. No difference was observed in the levels of AsA in the leaf and root tissues of the wild-type and knockout-nudx9 (KO-nudx9) plants, whereas a marked increase in N-glycoprotein levels and enhanced growth were detected in the roots of KO-nudx9 plants in the presence of ammonium. These results suggest that AtNUDX9 is involved in the regulation of GDP-d-Man levels affecting ammonium sensitivity via modulation of protein N-glycosylation in the roots.
Highlights
GDP-d-mannose (GDP-d-Man) is an activated sugar nucleotide that is required for many aspects of plant metabolism
Ogawa et al (2005) have already demonstrated that AtNUDX9 had no detectable activity toward various types of nucleotide sugars (ADP-d-ribose, ADP- and UDP-d-glucose, UDP-l-galactose), in addition to deoxynucleoside triphosphates, 8-oxo-7,8-dihydro2′-(deoxy) guanosine 5′-triphosphate, ApnA, NADH, FAD, and coenzyme A
It is possible that the AtNUDX9 reaction competes with that of VTC1 and perturbs the equilibration between intracellular d-Man 1-phosphate and the GDP-d-Man levels maintained by VTC1
Summary
GDP-d-mannose (GDP-d-Man) is an activated sugar nucleotide that is required for many aspects of plant metabolism This compound is necessary for the biosynthesis of ascorbic acid (AsA), an essential antioxidant in plants, and some different structural carbohydrates for the synthesis of cell wall polysaccharides, as a precursor (Bonin et al, 1997; Conklin et al, 1999) (see Supplementary Fig. S1 at JXB online). A mutation in the gene encoding enzymes downstream of VTC1 in the AsA biosynthetic pathway, l-galactose-1-phosphate phosphatase (GPP/ VTC4), whose products are only used to synthesize AsA (see Supplementary Fig. S1 at JXB online), hardly showed such pleiotropic phenotypes (Qin et al, 2008) It appears that some of these pleiotropic phenotypes of vtc cannot be directly attributed to an AsA deficiency, suggesting the importance of maintaining appropriate levels of GDP-d-Man in cells
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