Enzymatic Evidence for a Complete Oxidative Pentose Phosphate Pathway in Chloroplasts and an Incomplete Pathway in the Cytosol of Spinach Leaves

Schnarrenberger, Claus; Flechner, Anke; Martin, William J.

doi:10.1104/pp.108.2.609

Cited by 124 publications

(94 citation statements)

References 48 publications

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“…It also provides intermediates for other biosynthetic pathways ( Kruger and von Schaewen, 2003), for instance, for fatty acid biosynthesis, nitrate reduction, and the shikimate pathway (Emes and Neuhaus, 1997). In plants, it is compartmentalized within the cytosol and the plastids (Schnarrenberger et al, 1995). Both pathways are linked via a pentose phosphate translocator that is ubiquitously expressed in the various plant tissues (Eicks et al, 2002).…”

Section: Discussionmentioning

confidence: 99%

A Cytosolic Arabidopsis d-Xylulose Kinase Catalyzes the Phosphorylation of 1-Deoxy-d-Xylulose into a Precursor of the Plastidial Isoprenoid Pathway

et al. 2006

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Plants are able to integrate exogenous 1-deoxy-D-xylulose (DX) into the 2C-methyl-D-erythritol 4-phosphate pathway, implicated in the biosynthesis of plastidial isoprenoids. Thus, the carbohydrate needs to be phosphorylated into 1-deoxy-D-xylulose 5-phosphate and translocated into plastids, or vice versa. An enzyme capable of phosphorylating DX was partially purified from a cell-free Arabidopsis (Arabidopsis thaliana) protein extract. It was identified by mass spectrometry as a cytosolic protein bearing D-xylulose kinase (XK) signatures, already suggesting that DX is phosphorylated within the cytosol prior to translocation into the plastids. The corresponding cDNA was isolated and enzymatic properties of a recombinant protein were determined. In Arabidopsis, xylulose kinases are encoded by a small gene family, in which only two genes are putatively annotated. The additional gene is coding for a protein targeted to plastids, as was proved by colocalization experiments using green fluorescent protein fusion constructs. Functional complementation assays in an Escherichia coli strain deleted in XK revealed that the cytosolic enzyme could exclusively phosphorylate xylulose in vivo, not the enzyme that is targeted to plastids. XK activities could not be detected in chloroplast protein extracts or in proteins isolated from its ancestral relative Synechocystis sp. PCC 6803. The gene encoding the plastidic protein annotated as ''xylulose kinase'' might in fact yield an enzyme having different phosphorylation specificities. The biochemical characterization and complementation experiments with DX of specific Arabidopsis knockout mutants seedlings treated with oxo-clomazone, an inhibitor of 1-deoxy-D-xylulose 5-phosphate synthase, further confirmed that the cytosolic protein is responsible for the phosphorylation of DX in planta.In plants, the biosynthesis of the active isoprene units (D 2 -and D 3

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Section: Discussionmentioning

confidence: 99%

A Cytosolic Arabidopsis d-Xylulose Kinase Catalyzes the Phosphorylation of 1-Deoxy-d-Xylulose into a Precursor of the Plastidial Isoprenoid Pathway

et al. 2006

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“…In plants, TK is additionally required in the Calvin cycle, in which it converts glyceraldehyde-3-phosphate and Fru6P to xylulose-5-phosphate and Ery4P, and glyceraldehyde-3-phosphate and sedoheptulase-7-phosphate to ribose-5-phosphate and xylulose-5-phosphate. Plants contain one major isoform of TK (Schnarrenberger et al, 1995), which is located in the chloroplast (Schnarrenberger et al, 1995;Debnam and Emes, 1999). In spinach, TK encodes a protein with a plastid-targeting sequence (Teige et al, 1995;Flechner et al, 1996) and is expressed in photosynthetic and nonphotosynthetic tissues (Bernacchia et al, 1995;Teige et al, 1995), indicating that an amphibolic TK participates in the Calvin cycle and the plastid OPPP.…”

Section: Introductionmentioning

confidence: 99%

A Small Decrease of Plastid Transketolase Activity in Antisense Tobacco Transformants Has Dramatic Effects on Photosynthesis and Phenylpropanoid Metabolism

Henkes

Sonnewald²,

Badur³

et al. 2001

Plant Cell

303

156

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Transketolase (TK) catalyzes reactions in the Calvin cycle and the oxidative pentose phosphate pathway (OPPP) and produces erythrose-4-phosphate, which is a precursor for the shikimate pathway leading to phenylpropanoid metabolism. To investigate the consequences of decreased TK expression for primary and secondary metabolism, we transformed tobacco with a construct containing an antisense TK sequence. The results were as follows: (1) a 20 to 40% reduction of TK activity inhibited ribulose-1,5-bisphosphate regeneration and photosynthesis. The inhibition of photosynthesis became greater as irradiance increased across the range experienced in growth conditions (170 to 700 mol m Ϫ 2 sec Ϫ 1 ). TK almost completely limited the maximum rate of photosynthesis in saturating light and saturating CO 2 . (2) Decreased expression of TK led to a preferential decrease of sugars, whereas starch remained high until photosynthesis was strongly inhibited. One of the substrates of TK (fructose-6-phosphate) is the starting point for starch synthesis, and one of the products (erythrose-4-phosphate) inhibits phosphoglucose isomerase, which catalyzes the first reaction leading to starch. (3) A 20 to 50% decrease of TK activity led to decreased levels of aromatic amino acids and decreased levels of the intermediates (caffeic acid and hydroxycinnamic acids) and products (chlorogenic acid, tocopherol, and lignin) of phenylpropanoid metabolism. (4) There was local loss of chlorophyll and carotene on the midrib when TK activity was inhibited by Ͼ 50%, spreading onto minor veins and lamina in severely affected transformants. (5) OPPP activity was not strongly inhibited by decreased TK activity. These results identify TK activity as an important determinant of photosynthetic and phenylpropanoid metabolism and show that the provision of precursors by primary metabolism colimits flux into the shikimate pathway and phenylpropanoid metabolism. INTRODUCTIONThe interaction between primary and secondary pathways is an important but poorly understood aspect of plant metabolism. Phenylpropanoids represent an important class of secondary metabolites, with roles in plant structure, defense, and signaling (Dixon and Paiva, 1995). They are derived from aromatic amino acids, which are synthesized via the shikimate pathway in the plastid. Up to 20% of the total carbon in a plant passes through this pathway (Jensen, 1985). Even though there have been numerous studies of transformants with altered expression of genes that encode enzymes in the Calvin cycle, glycolysis, the tricarboxylic acid cycle, and the oxidative pentose phosphate pathway (OPPP) (Stitt and Sonnewald, 1995;Stitt, 1999), the consequences for phenylpropanoid metabolism or other secondary pathways have not been explored. The presence of redundant enzymes and pathways for carbohydrate breakdown and glycolysis (Dennis, 1987; Dennis et al., 1997;Stitt, 1998) and the absence of marked phenotypic changes after alterations in the expression of key enzymes, including phosphofructokinase (Burrell et...

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“…In plant cells, the complete OPPP operates in plastids (Schnarrenberger et al, 1995) and in chloroplasts at night as a formal reversion of the Calvin-Benson cycle (Buchanan, 1991;Scheibe, 1991;Kruger and von Schaewen, 2003). The three irreversible OPPP reactions also are present in the cytosol but probably only transiently in peroxisomes (Meyer et al, 2011;Hölscher et al, 2014).…”

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confidence: 99%

Defects in Peroxisomal 6-Phosphogluconate Dehydrogenase Isoform PGD2 Prevent Gametophytic Interaction in Arabidopsis thaliana

et al. 2016

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We studied the localization of 6-phosphogluconate dehydrogenase (PGD) isoforms of Arabidopsis (Arabidopsis thaliana). Similar polypeptide lengths of PGD1, PGD2, and PGD3 obscured which isoform may represent the cytosolic and/or plastidic enzyme plus whether PGD2 with a peroxisomal targeting motif also might target plastids. Reporter-fusion analyses in protoplasts revealed that, with a free N terminus, PGD1 and PGD3 accumulate in the cytosol and chloroplasts, whereas PGD2 remains in the cytosol. Mutagenesis of a conserved second ATG enhanced the plastidic localization of PGD1 and PGD3 but not PGD2. Amino-terminal deletions of PGD2 fusions with a free C terminus resulted in peroxisomal import after dimerization, and PGD2 could be immunodetected in purified peroxisomes. Repeated selfing of pgd2 transfer (T-)DNA alleles yielded no homozygous mutants, although siliques and seeds of heterozygous plants developed normally. Detailed analyses of the C-terminally truncated PGD2-1 protein showed that peroxisomal import and catalytic activity are abolished. Reciprocal backcrosses of pgd2-1 suggested that missing PGD activity in peroxisomes primarily affects the male gametophyte. Tetrad analyses in the quartet1-2 background revealed that pgd2-1 pollen is vital and in vitro germination normal, but pollen tube growth inside stylar tissues appeared less directed. Mutual gametophytic sterility was overcome by complementation with a genomic construct but not with a version lacking the first ATG. These analyses showed that peroxisomal PGD2 activity is required for guided growth of the male gametophytes and pollen tube-ovule interaction. Our report finally demonstrates an essential role of oxidative pentose-phosphate pathway reactions in peroxisomes, likely needed to sustain critical levels of nitric oxide and/or jasmonic acid, whose biosynthesis both depend on NADPH provision.

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Enzymatic Evidence for a Complete Oxidative Pentose Phosphate Pathway in Chloroplasts and an Incomplete Pathway in the Cytosol of Spinach Leaves

Cited by 124 publications

References 48 publications

A Cytosolic Arabidopsis d-Xylulose Kinase Catalyzes the Phosphorylation of 1-Deoxy-d-Xylulose into a Precursor of the Plastidial Isoprenoid Pathway

A Cytosolic Arabidopsis d-Xylulose Kinase Catalyzes the Phosphorylation of 1-Deoxy-d-Xylulose into a Precursor of the Plastidial Isoprenoid Pathway

A Small Decrease of Plastid Transketolase Activity in Antisense Tobacco Transformants Has Dramatic Effects on Photosynthesis and Phenylpropanoid Metabolism

Defects in Peroxisomal 6-Phosphogluconate Dehydrogenase Isoform PGD2 Prevent Gametophytic Interaction in Arabidopsis thaliana

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