Plant Science 166 (2004) 1335–1343 Characterization of a dicarboxylate exchange system able to exchange pyrophosphate for l-malate in non-photosynthetic plastids from developing maize embryos Aurora Lara-Núñez, Rogelio Rodr´ ıguez-Sotres ∗ Department of Biochemistry, Faculty of Chemistry, Universidad Nacional Autónoma de México, México City, Mexico Received 10 December 2003; received in revised form 20 January 2004; accepted 21 January 2004 Abstract Non-photosynthetic plastids from plant tissues are unable to fix carbon dioxide into carbohydrates, and so, they depend on carriers of the inner plastid membrane. Spinach chloroplasts are known to have a dicarboxylate translocator (DL) system able to exchange l-malate for other dicarboxylates. This paper presents evidence for the existence of a similar, but not equal system in non-photosynthetic plastids, isolated from developing maize embryos. Plastids were isolated by Percoll gradient centrifugation. Dicarboxylate uptake or release was determined by centrifugation through a single silicone layer. We found a dicarboxylate translocating activity able to exchange l-malate, for either succinate, glutamate, oxaloacetate, 2-oxoglutarate or aspartate, and also malonate and the tricarboxylate, citrate. Surprisingly, this activity also exchanged malate for pyrophosphate and in addition, pyrophosphate inhibited the dicarboxylate exchange. We found a competitive inhibition pattern of the l-malate/malonate exchange by n-butylmalonate, and this compound also inhibited the l-malate/pyrophosphate exchange. Our results are consistent with the participation of, at least, one common component in the l-malate/malonate and l-malate/pyrophosphate carrier systems. We discuss the possible role of the dicarboxylate exchange in the plastidial and cytoplasm metabolism in developing maize embryos. © 2004 Elsevier Ireland Ltd. All rights reserved. Keywords: Plastid; Dicarboxylate translocator; Pyrophosphate; Zea mays 1. Introduction Several translocators which reside on the inner membrane of the plastid envelope, coordinate the exchange of metabo- lites between the cytosol and the plastid stroma [1]. Plastids from developing embryos do not photosynthesize, but the metabolism of fatty acids, some amino acids and other important metabolites is active in their stroma [1]. Therefore, they depend on metabolite exchange with the cytosol to support all of these activities. Up to now, at Abbreviations: TPT, triose phosphate/phosphate translocator; HPT, hexose phosphate/phosphate translocator; PPT, phosphoenolpyruvate/ phosphate translocator; XPT, pentose-phosphate/phosphate translocator; DT, dicarboxylate translocator; HEPES, [4-(2-hydroxyethyl)-1-piperazi- neethanesulfonic acid]; NADP–ME, NADP-dependent malic enzyme; EDTA, ethylenediaminotetraacetic acid; DTT, dithiothreitol; Pi, phosphate; PPi, pyrophosphate ∗ Corresponding author. Present address: Ciudad Universitaria, Insur- gentes sur Copilco, Coyoac´ an M´ exico City, Mexico Tel.: +52-55-5622-5285; fax: +52-55-5622-5329. E-mail address: sotres@servidor.unam.mx (R. Rodr´ ıguez-Sotres). least six specific metabolite-exchange mediators have been identified in the envelope of non-photosynthetic plastids from several plant species: the triose phosphate/phosphate translocator (TPT) [2], the adenine nucleotides transloca- tor [3], the hexose phosphate/phosphate translocator (HPT) [4], the phosphoenolpyruvate/phosphate translocator (PPT) [5,6], the pentose-phosphate/phosphate translocator [7], and the malate/Pi translocator [8]. In the case of spinach leaf chloroplasts, a malate/oxalo- acetate shuttle may be required for the regulation of the re- dox balance, and a glutamate/malate and a 2-oxoglutarate/ malate shuttles could help to sustain stromal amino acid syn- thesis [9,10]. Both translocators participate in a 2-oxogluta- rate/glutamate exchange, with no net malate transport required during photorespiration [9,10]. Although the data presented by Taniguchi et al. [11] indicates that dicarboxy- late translocators (DT) are expressed in photosynthetic and non-photosynthetic tissues in Arabidopsis plants, the prop- erties of the system fulfilling such functions have not been documented in leucoplasts, or in other non-photosynthetic plastids. 0168-9452/$ – see front matter © 2004 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.plantsci.2004.01.015