title: Defects in Peroxisomal 6-Phosphogluconate Dehydrogenase Isoform PGD2 Prevent Gametophytic Interaction in Arabidopsis thaliana.
abstract: 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.
authors.affiliation: Institut für Biologie und Biotechnologie der Pflanzen, Westfälische Wilhelms-Universität Münster, Schlossplatz 7, D-48149 Münster, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Hölscher
authors.affiliation: Institut für Biologie und Biotechnologie der Pflanzen, Westfälische Wilhelms-Universität Münster, Schlossplatz 7, D-48149 Münster, Germany.
authors.forename: Marie-Christin
authors.identifier: 0000-0003-0869-1320
authors.initials: MC
authors.lastname: Lutterbey
authors.affiliation: Institut für Biologie und Biotechnologie der Pflanzen, Westfälische Wilhelms-Universität Münster, Schlossplatz 7, D-48149 Münster, Germany.
authors.forename: Hannes
authors.identifier: 0000-0001-5716-2877
authors.initials: H
authors.lastname: Lansing
authors.affiliation: Institut für Biologie und Biotechnologie der Pflanzen, Westfälische Wilhelms-Universität Münster, Schlossplatz 7, D-48149 Münster, Germany.
authors.forename: Tanja
authors.initials: T
authors.lastname: Meyer
authors.affiliation: Institut für Biologie und Biotechnologie der Pflanzen, Westfälische Wilhelms-Universität Münster, Schlossplatz 7, D-48149 Münster, Germany.
authors.forename: Kerstin
authors.initials: K
authors.lastname: Fischer
authors.affiliation: Institut für Biologie und Biotechnologie der Pflanzen, Westfälische Wilhelms-Universität Münster, Schlossplatz 7, D-48149 Münster, Germany schaewen@uni-muenster.de.
authors.forename: Antje
authors.identifier: 0000-0003-0832-108X
authors.initials: A
authors.lastname: von Schaewen
mesh_terms: D000483:Alleles; D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics; D003001:Cloning, Molecular; D003517:Cyclopentanes / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004269:DNA, Bacterial; D018744:DNA, Plant / Q000302:isolation & purification; D055993:Germ Cells, Plant / Q000187:drug effects*; D018525:Germination / Q000187:drug effects / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D009569:Nitric Oxide / Q000378:metabolism; D054883:Oxylipins / Q000378:metabolism; D010427:Pentose Phosphate Pathway; D020675:Peroxisomes / Q000378:metabolism; D010734:Phosphogluconate Dehydrogenase / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics; D018515:Plant Leaves / Q000378:metabolism; D018087:Plastids; D011058:Pollen / Q000187:drug effects / Q000254:growth & development; D015230:Prostaglandin D2 / Q000037:antagonists & inhibitors*; D011457:Prostaglandins D / Q000037:antagonists & inhibitors; D020033:Protein Isoforms / Q000037:antagonists & inhibitors*; D012639:Seeds / Q000187:drug effects / Q000254:growth & development; D020539:Sequence Analysis, Protein
chemical_list: D029681:Arabidopsis Proteins; D003517:Cyclopentanes; D004269:DNA, Bacterial; D018744:DNA, Plant; D054883:Oxylipins; D011457:Prostaglandins D; D020033:Protein Isoforms; C036483:T-DNA; D009569:Nitric Oxide; C011006:jasmonic acid; D010734:Phosphogluconate Dehydrogenase; D015230:Prostaglandin D2