Dataset Information

UDP-sugar pyrophosphorylase is essential for arabinose and xylose recycling, and is required during vegetative and reproductive growth in Arabidopsis.

ABSTRACT: Numerous nucleotide sugars are needed in plants to synthesize cell wall polymers and glycoproteins. The de novo synthesis of nucleotide sugars is of major importance. During growth, however, some polymers are broken down to monosaccharides. Reactivation of these sugars into nucleotide sugars occurs in two steps: first, by a substrate-specific sugar-1-kinase and, second, by UDP-sugar-pyrophosphorylase (USP), which has broad substrate specificity. A knock-out of the USP gene results in non-fertile pollen. By using various genetic complementation approaches we obtained a strong (>95%) knock-down line in USP that allowed us to investigate the physiological role of the enzyme during the life cycle. Mutant plants show an arabinose reduction in the cell wall, and accumulate mainly two sugars, arabinose and xylose, in the cytoplasm. The arabinogalactanproteins in usp mutants show no significant reduction in size. USP is also part of the myo-inositol oxygenation pathway to UDP-glucuronic acid; however, free glucuronic acid does not accumulate in cells, suggesting alternative conversion pathways of this monosaccharide. The knock-down plants are mostly sterile because of the improper formation of anthers and pollen sacks.

SUBMITTER: Geserick C

PROVIDER: S-EPMC3659416 | biostudies-literature | 2013 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

UDP-sugar pyrophosphorylase is essential for arabinose and xylose recycling, and is required during vegetative and reproductive growth in Arabidopsis.

Geserick Claudia C Tenhaken Raimund R

The Plant journal : for cell and molecular biology 20130213 2

Numerous nucleotide sugars are needed in plants to synthesize cell wall polymers and glycoproteins. The de novo synthesis of nucleotide sugars is of major importance. During growth, however, some polymers are broken down to monosaccharides. Reactivation of these sugars into nucleotide sugars occurs in two steps: first, by a substrate-specific sugar-1-kinase and, second, by UDP-sugar-pyrophosphorylase (USP), which has broad substrate specificity. A knock-out of the USP gene results in non-fertile ...[more]

PMID: 23373795

Similar Datasets

Project description:Leishmaniases are a set of tropical and sub-tropical diseases caused by protozoan parasites of the genus Leishmania whose severity ranges from self-healing cutaneous lesions to fatal visceral infections. Leishmania parasites synthesise a wide array of cell surface and secreted glycoconjugates that play important roles in infection. These glycoconjugates are particularly abundant in the promastigote form and known to be essential for establishment of infection in the insect midgut and effective transmission to the mammalian host. Since they are rich in galactose, their biosynthesis requires an ample supply of UDP-galactose. This nucleotide-sugar arises from epimerisation of UDP-glucose but also from an uncharacterised galactose salvage pathway. In this study, we evaluated the role of the newly characterised UDP-sugar pyrophosphorylase (USP) of Leishmania major in UDP-galactose biosynthesis. Upon deletion of the USP encoding gene, L. major lost the ability to synthesise UDP-galactose from galactose-1-phosphate but its ability to convert glucose-1-phosphate into UDP-glucose was fully maintained. Thus USP plays a role in UDP-galactose activation but does not significantly contribute to the de novo synthesis of UDP-glucose. Accordingly, USP was shown to be dispensable for growth and glycoconjugate biosynthesis under standard growth conditions. However, in a mutant seriously impaired in the de novo synthesis of UDP-galactose (due to deficiency of the UDP-glucose pyrophosphorylase) addition of extracellular galactose increased biosynthesis of the cell surface lipophosphoglycan. Thus under restrictive conditions, such as those encountered by Leishmania in its natural habitat, galactose salvage by USP may play a substantial role in biosynthesis of the UDP-galactose pool. We hypothesise that USP recycles galactose from the blood meal within the midgut of the insect for synthesis of the promastigote glycocalyx and thereby contributes to successful vector infection.

Project description:Glycosomes are peroxisome-related organelles of trypanosomatid parasites containing metabolic pathways, such as glycolysis and biosynthesis of sugar nucleotides, usually present in the cytosol of other eukaryotes. UDP-glucose pyrophosphorylase (UGP), the enzyme responsible for the synthesis of the sugar nucleotide UDP-glucose, is localized in the cytosol and glycosomes of the bloodstream and procyclic trypanosomes, despite the absence of any known peroxisome-targeting signal (PTS1 and PTS2). The questions that we address here are (i) is the unusual glycosomal biosynthetic pathway of sugar nucleotides functional and (ii) how is the PTS-free UGP imported into glycosomes? We showed that UGP is imported into glycosomes by piggybacking on the glycosomal PTS1-containing phosphoenolpyruvate carboxykinase (PEPCK) and identified the domains involved in the UGP/PEPCK interaction. Proximity ligation assays revealed that this interaction occurs in 3 to 10% of glycosomes, suggesting that these correspond to organelles competent for protein import. We also showed that UGP is essential for the growth of trypanosomes and that both the glycosomal and cytosolic metabolic pathways involving UGP are functional, since the lethality of the knockdown UGP mutant cell line (RNAiUGP, where RNAi indicates RNA interference) was rescued by expressing a recoded UGP (rUGP) in the organelle (RNAiUGP/EXPrUGP-GPDH, where GPDH is glycerol-3-phosphate dehydrogenase). Our conclusion was supported by targeted metabolomic analyses (ion chromatography-high-resolution mass spectrometry [IC-HRMS]) showing that UDP-glucose is no longer detectable in the RNAiUGP mutant, while it is still produced in cells expressing UGP exclusively in the cytosol (PEPCK null mutant) or glycosomes (RNAiUGP/EXPrUGP-GPDH). Trypanosomatids are the only known organisms to have selected functional peroxisomal (glycosomal) sugar nucleotide biosynthetic pathways in addition to the canonical cytosolic ones. IMPORTANCE Unusual compartmentalization of metabolic pathways within organelles is one of the most enigmatic features of trypanosomatids. These unicellular eukaryotes are the only organisms that sequestered glycolysis inside peroxisomes (glycosomes), although the selective advantage of this compartmentalization is still not clear. Trypanosomatids are also unique for the glycosomal localization of enzymes of the sugar nucleotide biosynthetic pathways, which are also present in the cytosol. Here, we showed that the cytosolic and glycosomal pathways are functional. As in all other eukaryotes, the cytosolic pathways feed glycosylation reactions; however, the role of the duplicated glycosomal pathways is currently unknown. We also showed that one of these enzymes (UGP) is imported into glycosomes by piggybacking on another glycosomal enzyme (PEPCK); they are not functionally related. The UGP/PEPCK association is unique since all piggybacking examples reported to date involve functionally related interacting partners, which broadens the possible combinations of carrier-cargo proteins being imported as hetero-oligomers.

Dataset Information

UDP-sugar pyrophosphorylase is essential for arabinose and xylose recycling, and is required during vegetative and reproductive growth in Arabidopsis.

Publications

UDP-sugar pyrophosphorylase is essential for arabinose and xylose recycling, and is required during vegetative and reproductive growth in Arabidopsis.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets