Screening of the Arabidopsis thaliana genome revealed three potential homologues of mammalian and yeast mitochondrial DICs (dicarboxylate carriers) designated as DIC1, DIC2 and DIC3, each belonging to the mitochondrial carrier protein family. DIC1 and DIC2 are broadly expressed at comparable levels in all the tissues investigated. DIC1-DIC3 have been reported previously as uncoupling proteins, but direct transport assays with recombinant and reconstituted DIC proteins clearly demonstrate that their substrate specificity is unique to plants, showing the combined characteristics of the DIC and oxaloacetate carrier in yeast. Indeed, the Arabidopsis DICs transported a wide range of dicarboxylic acids including malate, oxaloacetate and succinate as well as phosphate, sulfate and thiosulfate at high rates, whereas 2-oxoglutarate was revealed to be a very poor substrate. The role of these plant mitochondrial DICs is discussed with respect to other known mitochondrial carrier family members including uncoupling proteins. It is proposed that plant DICs constitute the membrane component of several metabolic processes including the malate-oxaloacetate shuttle, the most important redox connection between the mitochondria and the cytosol.

These authors contributed equally to this work. SummaryWe describe the identi®cation and functional characterization of two Arabidopsis mitochondrial basic amino acid carriers (BAC), AtmBAC1 and AtmBAC2, which are related to the yeast ornithine (Orn) carrier Ort1p, also known as Arg11p. The arg11 mutant requires arginine (Arg) supplementation because it fails to export suf®cient ornithine from the mitochondrion to the cytosol where it is converted to arginine. Atm-BAC1 and, to a lesser extent, AtmBAC2 partially replaced the function of Ort1p in yeast arg11. The more ef®cient putative carrier, AtmBAC1, was expressed in E. coli, puri®ed, and reconstituted into phospholipid vesicles, where it transported the basic L-amino acids arginine, lysine, ornithine and histidine (in order of decreasing af®nity). AtmBAC1 recognized L-histidine whereas both yeast Ort1p and the mammalian ortholog ORNT1p do not. Also different from ORNT1p, AtmBAC1 did not transport citrulline. AtmBAC1 appeared to be more stereospeci®c than the yeast and mammalian ornithine carriers, exhibiting greater preference for the L-forms of arginine, lysine and ornithine. By RT-PCR, both AtmBAC1 and AtmBAC2 transcripts were detected in stems, leaves,¯owers, siliques, and seedlings. Expression of AtmBAC1 in seedlings is consistent with its involvement in Arg breakdown in early seedling development, i.e. delivery of Arg to mitochondrial arginase. The K m (0.19 mM) for Arg uptake by AtmBAC1 was close to the value we previously determined for the saturable component of Arg uptake into intact mitochondria from soybean seedling cotyledons.

Despite much study of the role of S-adenosylmethionine (SAM) in the methylation of DNA, RNA, and proteins, and as a cofactor for a wide range of biosynthetic processes, little is known concerning the intracellular transport of this essential metabolite. Screening of the Arabidopsis (Arabidopsis thaliana) genome yielded two potential homologs of yeast (Saccharomyces cerevisiae) and human SAM transporters, designated as SAMC1 and SAMC2, both of which belong to the mitochondrial carrier protein family. The SAMC1 gene is broadly expressed at the organ level, although only in specialized tissues of roots with high rates of cell division, and appears to be up-regulated in response to wounding stress, whereas the SAMC2 gene is very poorly expressed in all organs/tissues analyzed. Direct transport assays with the recombinant and reconstituted SAMC1 were utilized to demonstrate that this protein displays a very narrow substrate specificity confined to SAM and its closest analogs. Further experiments revealed that SAMC1 was able to function in uniport and exchange reactions and characterized the transporter as highly active, but sensitive to physiologically relevant concentrations of S-adenosylhomocysteine, S-adenosylcysteine, and adenosylornithine. Green fluorescent protein-based cell biological analysis demonstrated targeting of SAMC1 to mitochondria. Previous proteomic analyses identified this protein also in the chloroplast inner envelope. In keeping with these results, bioinformatics predicted dual localization for SAMC1. These findings suggest that the provision of cytosolically synthesized SAM to mitochondria and possibly also to plastids is mediated by SAMC1 according to the relative demands for this metabolite in the organelles.