title: Plastid uridine salvage activity is required for photoassimilate allocation and partitioning in Arabidopsis.
abstract: Nucleotides are synthesized from de novo and salvage pathways. To characterize the uridine salvage pathway, two genes, UKL1 and UKL2, that tentatively encode uridine kinase (UK) and uracil phosphoribosyltransferase (UPRT) bifunctional enzymes were studied in Arabidopsis thaliana. T-DNA insertions in UKL1 and UKL2 reduced transcript expression and increased plant tolerance to toxic analogs 5-fluorouridine and 5-fluorouracil. Enzyme activity assays using purified recombinant proteins indicated that UKL1 and UKL2 have UK but not UPRT activity. Subcellular localization using a C-terminal enhanced yellow fluorescent protein fusion indicated that UKL1 and UKL2 localize to plastids. The ukl2 mutant shows reduced transient leaf starch during the day. External application of orotate rescued this phenotype in ukl2, indicating pyrimidine pools are limiting for starch synthesis in ukl2. Intermediates for lignin synthesis were upregulated, and there was increased lignin and reduced cellulose content in the ukl2 mutant. Levels of ATP, ADP, ADP-glucose, UTP, UDP, and UDP-glucose were altered in a light-dependent manner. Seed composition of the ukl1 and ukl2 mutants included lower oil and higher protein compared with the wild type. Unlike single gene mutants, the ukl1 ukl2 double mutant has severe developmental defects and reduced biomass accumulation, indicating these enzymes catalyze redundant reactions. These findings point to crucial roles played by uridine salvage for photoassimilate allocation and partitioning.
authors.affiliation: Division of Biochemistry and Interdisciplinary Plant Group, Christopher S. Bond Life Sciences Center, University of Missouri, Columbia, Missouri 65211, USA. chenmi@missouri.edu
authors.forename: Mingjie
authors.initials: M
authors.lastname: Chen
authors.forename: Jay J
authors.initials: JJ
authors.lastname: Thelen
mesh_terms: D017360:Arabidopsis / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics / Q000502:physiology; D029681:Arabidopsis Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D018533:Biomass; D002482:Cellulose / Q000378:metabolism; D005472:Fluorouracil / Q000494:pharmacology; D007527:Isoenzymes / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008027:Light; D008031:Lignin / Q000378:metabolism; D016254:Mutagenesis, Insertional; D009963:Orotic Acid / Q000494:pharmacology; D010430:Pentosyltransferases / Q000235:genetics / Q000378:metabolism; D010641:Phenotype; D018515:Plant Leaves / Q000187:drug effects / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D030821:Plants, Genetically Modified; D018087:Plastids / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics / Q000502:physiology; D011993:Recombinant Fusion Proteins; D012639:Seeds / Q000187:drug effects / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D013213:Starch / Q000378:metabolism; D014498:Uracil / Q000494:pharmacology; D014529:Uridine / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D014541:Uridine Kinase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D029681:Arabidopsis Proteins; D007527:Isoenzymes; D011993:Recombinant Fusion Proteins; C001943:5-fluorouridine; D014498:Uracil; D009963:Orotic Acid; D002482:Cellulose; D013213:Starch; D008031:Lignin; D010430:Pentosyltransferases; C023269:uracil phosphoribosyltransferase; D014541:Uridine Kinase; D005472:Fluorouracil; D014529:Uridine