RRC ID 35455
Author Witz S, Jung B, Fürst S, Möhlmann T.
Title De novo pyrimidine nucleotide synthesis mainly occurs outside of plastids, but a previously undiscovered nucleobase importer provides substrates for the essential salvage pathway in Arabidopsis.
Journal Plant Cell
Abstract Nucleotide de novo synthesis is highly conserved among organisms and represents an essential biochemical pathway. In plants, the two initial enzymatic reactions of de novo pyrimidine synthesis occur in the plastids. By use of green fluorescent protein fusions, clear support is provided for a localization of the remaining reactions in the cytosol and mitochondria. This implies that carbamoyl aspartate, an intermediate of this pathway, must be exported and precursors of pyrimidine salvage (i.e., nucleobases or nucleosides) are imported into plastids. A corresponding uracil transport activity could be measured in intact plastids isolated from cauliflower (Brassica oleracea) buds. PLUTO (for plastidic nucleobase transporter) was identified as a member of the Nucleobase:Cation-Symporter1 protein family from Arabidopsis thaliana, capable of transporting purine and pyrimidine nucleobases. A PLUTO green fluorescent protein fusion was shown to reside in the plastid envelope after expression in Arabidopsis protoplasts. Heterologous expression of PLUTO in an Escherichia coli mutant lacking the bacterial uracil permease uraA allowed a detailed biochemical characterization. PLUTO transports uracil, adenine, and guanine with apparent affinities of 16.4, 0.4, and 6.3 μM, respectively. Transport was markedly inhibited by low concentrations of a proton uncoupler, indicating that PLUTO functions as a proton-substrate symporter. Thus, a protein for the absolutely required import of pyrimidine nucleobases into plastids was identified.
Volume 24(4)
Pages 1549-59
Published 2012-4
DOI 10.1105/tpc.112.096743
PII tpc.112.096743
PMID 22474184
PMC PMC3398563
MeSH Adenine / metabolism Amino Acid Sequence Arabidopsis / drug effects Arabidopsis / genetics Arabidopsis / metabolism* Arabidopsis Proteins / chemistry Arabidopsis Proteins / genetics Arabidopsis Proteins / metabolism* Biosynthetic Pathways* / drug effects Brassica / metabolism Carbonyl Cyanide m-Chlorophenyl Hydrazone / pharmacology Gene Expression Regulation, Plant / drug effects Green Fluorescent Proteins / metabolism Guanine / metabolism Intracellular Membranes / drug effects Intracellular Membranes / metabolism Models, Biological Molecular Sequence Data Nucleobase Transport Proteins / chemistry Nucleobase Transport Proteins / genetics Nucleobase Transport Proteins / metabolism* Nucleotides / biosynthesis* Plastids / drug effects Plastids / metabolism* Protein Transport / drug effects Protoplasts / drug effects Protoplasts / metabolism Pyrimidines / biosynthesis* RNA, Messenger / genetics RNA, Messenger / metabolism Recombinant Fusion Proteins / metabolism Sequence Alignment Sodium Chloride / pharmacology Substrate Specificity / drug effects Symporters / chemistry Symporters / genetics Symporters / metabolism* Time Factors Uracil / metabolism
IF 8.631
Times Cited 33
WOS Category PLANT SCIENCES BIOCHEMISTRY & MOLECULAR BIOLOGY CELL BIOLOGY
Resource
Prokaryotes E. coli JD23420(uraA)