In many cells and specially in muscle, mitochondria form elongated filaments or a branched reticulum. We show that Mfn2 (mitofusin 2), a mitochondrial membrane protein that participates in mitochondrial fusion in mammalian cells, is induced during myogenesis and contributes to the maintenance and operation of the mitochondrial network. Repression of Mfn2 caused morphological and functional fragmentation of the mitochondrial network into independent clusters. Concomitantly, repression of Mfn2 reduced glucose oxidation, mitochondrial membrane potential, cell respiration, and mitochondrial proton leak. We also show that the Mfn2-dependent mechanism of mitochondrial control is disturbed in obesity by reduced Mfn2 expression. In all, our data indicate that Mfn2 expression is crucial in mitochondrial metabolism through the maintenance of the mitochondrial network architecture, and reduced Mfn2 expression may explain some of the metabolic alterations associated with obesity.

We have identified a new human cDNA, L-amino acid transporter-2 (LAT-2), that induces a system L transport activity with 4F2hc (the heavy chain of the surface antigen 4F2, also named CD98) in oocytes. Human LAT-2 is the fourth member of the family of amino acid transporters that are subunits of 4F2hc. The amino acid transport activity induced by the co-expression of 4F2hc and LAT-2 was sodium-independent and showed broad specificity for small and large zwitterionic amino acids, as well as bulky analogs (e.g. BCH (2-aminobicyclo-(2,2,1)-heptane-2-carboxylic acid)). This transport activity was highly trans-stimulated, suggesting an exchanger mechanism of transport. Expression of tagged N-myc-LAT-2 alone in oocytes did not induce amino acid transport, and the protein had an intracellular location. Co-expression of N-myc-LAT-2 and 4F2hc gave amino acid transport induction and expression of N-myc-LAT-2 at the plasma membrane of the oocytes. These data suggest that LAT-2 is an additional member of the family of 4F2 light chain subunits, which associates with 4F2hc to express a system L transport activity with broad specificity for zwitterionic amino acids. Human LAT-2 mRNA is expressed in kidney >>> placenta > > brain, liver > spleen, skeletal muscle, heart, small intestine, and lung. Human LAT-2 gene localizes at chromosome 14q11.2-13 (13 cR or ϳ286 kb from marker D14S1349). The high expression of LAT-2 mRNA in epithelial cells of proximal tubules, the basolateral location of 4F2hc in these cells, and the amino acid transport activity of LAT-2 suggest that this transporter contributes to the renal reabsorption of neutral amino acids in the basolateral domain of epithelial proximal tubule cells.Last year, three amino acid transporter cDNAs (LAT-1, y ϩ LAT-1, and y ϩ LAT-2) 1 were identified as subunits of the heavy chain of the cell surface antigen 4F2 (4F2hc, also named CD98) (1-3). These subunits co-express amino acid transport activity with 4F2hc in oocytes (i.e. system L for LAT-1, and system y ϩ L for y ϩ LAT-1 and y ϩ LAT-2) (1-4). The role of this family of proteins in amino acid transport has recently been demonstrated by the fact that mutations in the y ϩ LAT-1 gene cause lysinuric protein intolerance, an inherited amino aciduria due to a defective renal reabsorption mechanism of dibasic amino acids (5, 6). The structural and functional similarities between 4F2hc and its homologous protein rBAT suggest that a member of this family of subunits might be the subunit of rBAT needed to fully express the amino acid transport system b o,ϩ activity (reviewed in Refs. 7 and 8). After the identification of rBAT as the Type I cystinuria gene (9), this subunit is a good candidate for non-Type I cystinuria (7). A search throughout gene data bases suggests that there may be as many as four new human members of the family of subunits of 4F2hc and rBAT.Kanai and co-workers (1) identified rat LAT-1 (also known as TA1) by co-expression cloning with 4F2hc in oocytes. The coexpressed transport activity shows clear characteristic...

Mitofusin-2 (Mfn2) is a mitochondrial membrane protein that participates in mitochondrial fusion in mammalian cells and mutations in the Mfn2 gene cause Charcot-Marie-Tooth neuropathy type 2A. Here, we show that Mfn2 loss-of-function inhibits pyruvate, glucose and fatty acid oxidation and reduces mitochondrial membrane potential, whereas Mfn2 gain-of-function increases glucose oxidation and mitochondrial membrane potential. As to the mechanisms involved, we have found that Mfn2 loss-of-function represses nuclear-encoded subunits of OXPHOS complexes I, II, III and V, whereas Mfn2 overexpression induced the subunits of complexes I, IV and V. Obesity-induced Mfn2 deficiency in rat skeletal muscle was also associated with a decrease in the subunits of complexes I, II, III and V. In addition, the effect of Mfn2 overexpression on mitochondrial metabolism was mimicked by a truncated Mfn2 mutant that is inactive as a mitochondrial fusion protein. Our results indicate that Mfn2 triggers mitochondrial energization, at least in part, by regulating OXPHOS expression through signals that are independent of its role as a mitochondrial fusion protein.

Cystinuria (MIM 220100) is a common recessive disorder of renal reabsorption of cystine and dibasic amino acids. Mutations in SLC3A1, encoding rBAT, cause cystinuria type I (ref. 1), but not other types of cystinuria (ref. 2). A gene whose mutation causes non-type I cystinuria has been mapped by linkage analysis to 19q12-13.1 (Refs 3,4). We have identified a new transcript, encoding a protein (bo, +AT, for bo,+ amino acid transporter) belonging to a family of light subunits of amino acid transporters, expressed in kidney, liver, small intestine and placenta, and localized its gene (SLC7A9) to the non-type I cystinuria 19q locus. Co-transfection of bo,+AT and rBAT brings the latter to the plasma membrane, and results in the uptake of L-arginine in COS cells. We have found SLC7A9 mutations in Libyan-Jews, North American, Italian and Spanish non-type I cystinuria patients. The Libyan Jewish patients are homozygous for a founder missense mutation (V170M) that abolishes b o,+AT amino-acid uptake activity when co-transfected with rBAT in COS cells. We identified four missense mutations (G105R, A182T, G195R and G295R) and two frameshift (520insT and 596delTG) mutations in other patients. Our data establish that mutations in SLC7A9 cause non-type I cystinuria, and suggest that bo,+AT is the light subunit of rBAT.

Caveolins (CAV) are essential components of caveolae; plasma membrane invaginations with reduced fluidity, reflecting cholesterol accumulation [1]. CAV proteins bind cholesterol, and CAV’s ability to move between cellular compartments helps control intracellular cholesterol fluxes [1–3]. In humans, CAV1 mutations result in lipodystrophy, cell transformation, and cancer [4–7]. CAV1 gene-disrupted mice exhibit cardiovascular diseases, diabetes, cancer, atherosclerosis, and pulmonary fibrosis [8, 9]. The mechanism(s) underlying these disparate effects are unknown, but our past work suggested CAV1 deficiency might alter metabolism: CAV1−/− mice exhibit impaired liver regeneration unless supplemented with glucose, suggesting systemic inefficiencies requiring additional metabolic intermediates [10]. Establishing a functional link between CAV1 and metabolism would provide a unifying theme to explain these myriad pathologies [11]. Here, we demonstrate that impaired proliferation and low survival with glucose restriction is a shortcoming of CAV1 deficient cells, caused by impaired mitochondrial function. Without CAV1, free cholesterol accumulates in mitochondrial membranes, increasing membrane condensation and reducing efficiency of the respiratory chain and intrinsic anti-oxidant defence. Upon activation of oxidative phosphorylation, this promotes accumulation of reactive oxygen species resulting in cell death. We confirm that this mitochondrial dysfunction predisposes CAV1 deficient animals to mitochondrial related diseases such as steatohepatitis and neurodegeneration.