The mitochondrial dicarboxylate carrier is essential for the growth of Saccharomyces cerevisiae on ethanol or acetate as the sole carbon source

Abstract

SummaryThe dicarboxylate carrier (DIC) is an integral membrane protein that catalyses a dicarboxylate-phosphate exchange across the inner mitochondrial membrane. We generated a yeast mutant lacking the gene for the DIC. The deletion mutant failed to grow on acetate or ethanol as sole carbon source but was viable on glucose, galactose, pyruvate, lactate and glycerol. The growth on ethanol or acetate was largely restored by the addition of low concentrations of aspartate, glutamate, fumarate, citrate, oxoglutarate, oxaloacetate and glucose, but not of succinate, leucine and lysine. The expression of the DIC gene in wild-type yeast was repressed in media containing ethanol or acetate with or without glycerol. These results indicate that the primary function of DIC is to transport cytoplasmic dicarboxylates into the mitochondrial matrix rather than to direct carbon flux to gluconeogenesis by exporting malate from the mitochondria. The ⌬DIC mutant may serve as a convenient host for overexpression of DIC and for the demonstration of its correct targeting and assembly.

The Channel-Forming Sym1 Protein Is Transported by the TIM23 Complex in a Presequence-Independent Manner

“…A common feature of carrier proteins in the inner membrane is their organization into high-molecular-mass complexes, which can be analyzed by BN-PAGE (42,43). To assess complex formation, Sym1 ZZ mitochondria were solubilized, and the complexes were analyzed by BN-PAGE and Western blotting.…”

The Ustilago maydis Nit2 Homolog Regulates Nitrogen Utilization and Is Required for Efficient Induction of Filamentous Growth

“…These are upregulated under nitrogen starvation but seem to be repressed by Nit2. While the closest yeast homolog of Mct1 is the riboflavin transporter Mch5p (68), yeast Dic1p represents a mitochondrial carboxylate transporter (43) that is thought to be required for providing carbon backbones to the mitochondrial matrix (64). The fact that Asp and Glu supplementation restores growth on ethanol or acetate in a yeast dic1 deletion strains indicates potential cross-talk between carbon and nitrogen metabolism via UmDic1 (64).…”

Identification of the Yeast Mitochondrial Transporter for Oxaloacetate and Sulfate

“…Molecular ammonia is protonated inside the mitochondria, and the pH gradient collapses. The rate of swelling of mitochondria was monitored by the decrease in A 546 (7). Yeast mitochondria (100 g of protein) were added to 1 ml of various ammonium salts (120 mM), 20 mM Tris-HCl, pH 7.4, 1 mM EDTA, 5 M rotenone, and 0.1 M antimycin.…”

“…S. cerevisiae encodes 35 members of the mitochondrial carrier family (1)(2)(3)(4), including the dicarboxylate and succinate-fumarate carriers (5,6). The former catalyzes the import into mitochondria of succinate (or malate) in exchange for phosphate, producing a net uptake of succinate and supply of a substrate to the Krebs cycle (7). The latter exchanges external succinate for fumarate and is required for gluconeogenesis (6,8).…”

“…Yeast gene disruption strains (⌬-DIC1 and ⌬-SFC1) cannot grow on either ethanol or acetate as sole carbon source, but they grow well on glycerol, pyruvate and other nonfermentable substrates (7,9). Growth of the ⌬-DIC1 strain on ethanol or acetate is restored by both low concentrations of oxaloacetate and other compounds that start the tricarboxylate cycle (and the oxidation of the acetyl-CoA unconsumed by the glyoxylate cycle) by generating either intramitochondrial oxaloacetate or other Krebs cycle intermediates (7). Therefore, does oxaloacetate enter yeast mitochondria, and if so, how?…”