Tetrahydrofolate biosynthesis in plants: molecular and functional characterization of dihydrofolate synthetase and three isoforms of folylpolyglutamate synthetase in Arabidopsis thaliana.
Tetrahydrofolate coenzymes involved in one-carbon (C1) metabolism are polyglutamylated. In organisms that synthesize tetrahydrofolate de novo, dihydrofolate synthetase (DHFS) and folylpolyglutamate synthetase (FPGS) catalyze the attachment of glutamate residues to the folate molecule. In this study we isolated cDNAs coding a DHFS and three isoforms of FPGS from Arabidopsis thaliana. The function of each enzyme was demonstrated by complementation of yeast mutants deficient in DHFS or FPGS activity, and by measuring in vitro glutamate incorporation into dihydrofolate or tetrahydrofolate. DHFS is present exclusively in the mitochondria, making this compartment the sole site of synthesis of dihydrofolate in the plant cell. In contrast, FPGS is present as distinct isoforms in the mitochondria, the cytosol, and the chloroplast. Each isoform is encoded by a separate gene, a situation that is unique among eukaryotes. The compartmentation of FPGS isoforms is in agreement with the predominance of gamma-glutamyl-conjugated tetrahydrofolate derivatives and the presence of serine hydroxymethyltransferase and C1-tetrahydrofolate interconverting enzymes in the cytosol, the mitochondria, and the plastids. Thus, the combination of FPGS with these folate-mediated reactions can supply each compartment with the polyglutamylated folate coenzymes required for the reactions of C1 metabolism. Also, the multicompartmentation of FPGS in the plant cell suggests that the transported forms of folate are unconjugated.
Ravanel
S
S
Laboratoire de Physiologie Cellulaire Végétale, Unité Mixte de Recherche 5019, Centre National de la Recherche Scientifique/Commissariat à l'Energie Atomique/Université Joseph Fourier, 38054 Grenoble Cedex 9, France. sravanel@cea.fr
Cherest
H
H
Jabrin
S
S
Grunwald
D
D
Surdin-Kerjan
Y
Y
Douce
R
R
Rébeillé
F
F
D017360:Arabidopsis / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D018076:DNA, Complementary; D005816:Genetic Complementation Test; D007527:Isoenzymes / Q000378:metabolism*; D008969:Molecular Sequence Data; D010453:Peptide Synthases / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny; D012441:Saccharomyces cerevisiae / Q000235:genetics; D013347:Subcellular Fractions / Q000201:enzymology; D013763:Tetrahydrofolates / Q000096:biosynthesis*
D018076:DNA, Complementary; D007527:Isoenzymes; D013763:Tetrahydrofolates; C030371:5,6,7,8-tetrahydrofolic acid; D010453:Peptide Synthases; C020041:dihydrofolate synthetase; C023103:folylpolyglutamate synthetase