A fluoroorotic acid-resistant mutant of Arabidopsis defective in the uptake of uracil

Abstract

A fluoroorotic acid (FOA)-resistant mutant of Arabidopsis thaliana was isolated by screening M2 populations of ethyl methane sulphonate (EMS)-mutagenized Columbia seed. FOA resistance was due to a nuclear recessive gene, for1-1, which locates to a 519 kb region in chromosome 5. Assays of key regulatory enzymes in de novo pyrimidine synthesis (uridine monophosphate synthase) and salvage biochemistry (thymidine kinase) confirmed that FOA resistance in for1-1/for1-1 plants was not due to altered enzymatic activities. Uptake studies using radiolabelled purines, pyrimidines, and [14C]FOA reveal that for1-1/for1-1 plants were specifically defective in the uptake of uracil or uracil-like bases. To confirm such specificity, genetic crosses show that FOR1 is a distinct locus from FUR1 which encodes a deoxyuridine nucleoside transporter. In addition, for1-1/for1-1 plants were restored to FOA sensitivity by transformation with the Escherichia coli uracil transporter gene uraA driven by the cauliflower mosaic virus (CaMV) 35S promoter. Molecular mapping studies reveal that FOR1 does not correspond to loci belonging to any of the six known nucleobase transporter families identified in the Arabidopsis genome. Moreover, FOR1 does not appear to regulate the transcript levels of either uracil transporter-encoding loci At2g03590 or At2g03530. The above results strongly suggest that the for1-1 mutant allele affects a transport mechanism that is specific for the uptake of uracil.

Uric acid in plants and microorganisms: Biological applications and genetics - A review

“…Studies using radiolabelled purins, pirimidines and [ 14 C] fluoroorotic acid revealed that the FOA recessive genes for “1-1/for 1-1” on chromosome 5 were unable to uptake uracil or uracil-like bases in Arabidopsis thaliana mutant [123].…”

“…To date, six loci along chromosome 5 of Arabidopsis genome were identified to encode nucleobase transporters: At5g03555 (from PRT family); At5g25420, At5g49990, and At5g62890 (from NAT family); At5g50300 (an AzgA-like transporter); and At5g41160 (from PUP family) [123], [124]. The recently characterized AzgA adenine–guanine–hypoxanthine transporter of Aspergillus nidulans was found to have amino acid similarity to Arabidopsis loci At5g50300 and At3g10960 encode proteins [125].…”

“…The recently characterized AzgA adenine–guanine–hypoxanthine transporter of Aspergillus nidulans was found to have amino acid similarity to Arabidopsis loci At5g50300 and At3g10960 encode proteins [125]. The amino acid sequence of the FUR4 uracil transporter of Saccharomyces cerevisiae (from PRT family) showed significant similarity to that of Arabidopsis locus At5g03555 encoded protein [123].…”

Modeling, Substrate Docking, and Mutational Analysis Identify Residues Essential for the Function and Specificity of a Eukaryotic Purine-Cytosine NCS1 Transporter

“…Most cells in all domains of life possess specific nucleobase transporters, distinct from those involved in the uptake of nucleosides (1,2). Nucleobase transporters are important in protozoan and plant development and in the susceptibility of human, protozoan, or fungal cells to purine-related drugs (3)(4)(5)(6)(7)(8)(9)(10). Fungi possess three evolutionary distinct families of transporters specific for nucleobases (2,11,12).…”

Nucleoside transport and associated metabolism*

“…In a screen for fluoroorotic acid (FOA)‐resistant plants, the for1 mutant was identified. Uptake studies using radiolabelled purines, pyrimidines and [ 14 C]‐FOA reveal that for1 plants were specifically defective in the uptake of uracil or uracil‐like bases (Mourad et al. 2006).…”