PerspectiveCell Biology

A Mitochondrial Mystery, Solved

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Science  06 Jul 2012:
Vol. 337, Issue 6090, pp. 41-43
DOI: 10.1126/science.1225601

The three-carbon molecule pyruvate is a metabolic intermediate and a central hub for cellular energy metabolism. It lies at the junction of aerobic and anaerobic metabolism and is the precursor for biosynthetic pathways including glucose, lipid, and amino acid syntheses. Given the ubiquitous role of pyruvate in cellular bioenergetics (see the figure), its subcellular localization plays a critical role in its fate. The final product of pyruvate metabolism is fundamentally altered once it enters the mitochondrion. As such, the identity of the protein that transports pyruvate from the cytoplasm into mitochondria has been eagerly anticipated, and the wait has been nearly 40 years. Two papers in this issue, by Bricker et al. (1) on page 96 and Herzig et al. on page 93 (2), report the identification of a mitochondrial pyruvate carrier (MPC) responsible for this function—a momentous development in the field of bioenergetics with profound implications for treating metabolic diseases.

Although pyruvate, a weak acid, can passively cross the mitochondrial inner membrane, extensive biochemical characterization demonstrated the specificity and kinetics of its uptake by mitochondria (38) (the outer membrane is not believed to provide a significant barrier as it is permeable to small solutes, including pyruvate). The discovery that thiol-reactive agents, such as α-cyano-4-hydroxycinnamate, inhibit pyruvate transport across both the mitochondrial and plasma membranes (3, 5) firmly established facilitated transport of pyruvate.

But the identity of the pyruvate carrier had been a long-standing mystery. Bricker et al. used an elegant combination of yeast genetics, metabolomics, and mutational analysis of cells from patients with defects in mitochondrial pyruvate metabolism to identify two MPC family members (MPC1 and MPC2), each ∼15 kD in size, obligatory for mitochondrial pyruvate transport in yeast, fruit flies, and mammals. MPC is likely a multimeric complex in the mitochondrial inner membrane, and deletions in genes encoding MPC components caused deficient carbohydrate metabolism in the fruit fly Drosophila melanogaster and in yeast; the latter also became auxotrophic for leucine. Metabolomics pinpointed MPC1 as facilitating the conversion of cytosolic pyruvate to mitochondrial acetyl–coenzyme A (CoA). Functional analysis with the high-affinity MPC inhibitor UK5099 (5) implicated MPC1 as a crucial mediator of pyruvate transport activity. Unequivocal proof, however, requires the reconstitution of inhibitor-sensitive transport activity in a system lacking MPC1 and MPC2.

Paths of pyruvate.

Pyruvate can either passively diffuse across the mitochondrial inner membrane or be transported by the mitochondrial pyruvate carrier (MPC); both processes are driven by ΔpH. MCT, monocarboxylate transporter; GLUT, glucose transporter; TCA, tricarboxylic acid; NADH, nicotinamide adenine dinucleotide, reduced; FADH2, flavin adenine dinucleotide, reduced; ATP, adenosine-5′-triphosphate; CHC, cinnamic acids.

CREDIT: Y. HAMMOND/SCIENCE

Such proof is provided by Herzig et al. In addition to showing defective lipoic acid biosynthesis and impaired pyruvate uptake in yeast mitochondria lacking MPC paralogs, the authors demonstrated that MPC1 and MCP2 are sufficient to facilitate pyruvate uptake. When expressed in the lactic acid bacteria Lactococcus lactis, which maintain a steady-state pH gradient (ΔpH) comparable to that in isolated mitochondria, neither MPC1 nor MPC2 showed appreciable transport activity. Coexpression of both proteins, however, was sufficient to reconstitute pyruvate uptake. The transport was inhibited by UK5099 and modulated by manipulation of ΔpH, two hallmark characteristics of the long-sought mitochondrial pyruvate carrier.

Why has the identity of the mitochondrial pyruvate transporter remained so elusive? Curiously, the components of this transporter may have been purified as early as 1981 (9), though subsequent efforts to identify the proteins proved difficult (10). Perhaps the greatest impediment was assumptions about its structure. Although the shared pharmacology of inhibiting pyruvate transport with α-cyano-4-hydroxycinnamate at the plasma and mitochondrial membranes hinted that the transporters might be related, transport at the plasma membrane was later discovered to be facilitated by monocarboxylate transporters (MCTs, the SLC16 gene family, ∼35 to 50 kD) with inhibitor and substrate specificities distinct from those of the mitochondrial pyruvate carrier (11). Another assumption was that the transporter should be a member of the mitochondrial carrier protein family (SLC25 gene family, ∼28 to 34 kD) (12), which transports nucleotides, anions, and carboxylic acids involved in mitochondrial energy metabolism such as malate and citrate. As much of this family has yet to be functionally characterized, it seemed safe to propose that the mitochondrial pyruvate transporter would be a member.

The composition of the MPC complex is therefore extraordinary. The functional reconstitution experiments of Herzig et al. showed that neither MPC1 nor MPC2 in isolation can function as a monomer (unlike mitochondrial carrier proteins), but transport requires both. Moreover, immunoprecipitation and native gel electrophoresis conducted by Bricker et al. suggest that the MPC is a large heterocomplex, potentially with unequal stoichiometry between the two proteins. Such a structure is a fundamentally different paradigm for mitochondrial substrate transport and studies of its regulation could prove fascinating.

Given that the mitochondrial pyruvate transporter regulates a crucial branch point in cellular metabolism, its identification has enormous potential to treat human disease. One aspect is highlighted by Bricker et al., who link mutations in MPC1 to families with a devastating multisystem genetic disease caused by impaired mitochondrial pyruvate oxidation. Sequencing of the MPC1 and MPC2 genes in the diagnosis of mitochondrial disorders may help tailor appropriate choices of dietary and therapeutic intervention for these patients.

More broadly, knowledge of the components of a pyruvate transporter may facilitate emerging drug discovery efforts targeted to mitochondria (13). Agents that enhance the rate of mitochondrial pyruvate uptake might reverse the Warburg effect, a hallmark of cancer in which tumor cells rely heavily on glycolytic metabolism despite adequate oxygen supply. Indeed, enhancing the rate of mitochondrial pyruvate oxidation with dichloroacetate (DCA) is being clinically evaluated for the treatment of cancer (14).

Although a systematic review of DCA as a treatment for mitochondrial disorders has not indicated widespread success (15), enhanced mitochondrial pyruvate transport and oxidation could prove effective for the subset of patients for whom deficient pyruvate uptake and oxidation are causative of disease. Furthermore, the extent to which pyruvate transport controls the rate of oxidation has perhaps been underestimated in some tissues (9), so targeting the MPC may provide avenues for treatment where DCA has failed. Identification of the MPC also has implications for treating other pathologies, including heart failure, ischemia/reperfusion injury, and type 2 diabetes. Modulation of pyruvate transport could potentiate metabolic flexibility and respiratory capacity that might be beneficial to treating these diseases.

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