The role of mitochondria in cell metabolism and survival is controlled by calcium signals that are commonly transmitted at the close associations between mitochondria and endoplasmic reticulum (ER). However, the physical linkage of the ER–mitochondria interface and its relevance for cell function remains elusive. We show by electron tomography that ER and mitochondria are adjoined by tethers that are ∼10 nm at the smooth ER and ∼25 nm at the rough ER. Limited proteolysis separates ER from mitochondria, whereas expression of a short “synthetic linker” (<5 nm) leads to tightening of the associations. Although normal connections are necessary and sufficient for proper propagation of ER-derived calcium signals to the mitochondria, tightened connections, synthetic or naturally observed under apoptosis-inducing conditions, make mitochondria prone to Ca2+ overloading and ensuing permeability transition. These results reveal an unexpected dependence of cell function and survival on the maintenance of proper spacing between the ER and mitochondria.

Neuromuscular diseases are often caused by inherited mutations that lead to progressive skeletal muscle weakness and degeneration. In diverse populations of normal healthy mice, we observed correlations between the abundance of mRNA transcripts related to mitochondrial biogenesis, the dystrophin-sarcoglycan complex, and nicotinamide adenine dinucleotide (NAD+) synthesis, consistent with a potential role for the essential cofactor NAD+ in protecting muscle from metabolic and structural degeneration. Furthermore, the skeletal muscle transcriptomes of patients with Duchene’s muscular dystrophy (DMD) and other muscle diseases were enriched for various poly[adenosine 5’-diphosphate (ADP)–ribose] polymerases (PARPs) and for nicotinamide N-methyltransferase (NNMT), enzymes that are major consumers of NAD+ and are involved in pleiotropic events, including inflammation. In the mdx mouse model of DMD, we observed significant reductions in muscle NAD+ levels, concurrent increases in PARP activity, and reduced expression of nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme for NAD+ biosynthesis. Replenishing NAD+ stores with dietary nicotinamide riboside supplementation improved muscle function and heart pathology in mdx and mdx/Utr−/− mice and reversed pathology in Caenorhabditis elegans models of DMD. The effects of NAD+ repletion in mdx mice relied on the improvement in mitochondrial function and structural protein expression (α-dystrobrevin and δ-sarcoglycan) and on the reductions in general poly(ADP)-ribosylation, inflammation, and fibrosis. In combination, these studies suggest that the replenishment of NAD+ may benefit patients with muscular dystrophies or other neuromuscular degenerative conditions characterized by the PARP/NNMT gene expression signatures.

The BH3-only BID (BH3-interacting domain death agonist) protein has a critical function in the death-receptor pathway in the liver by triggering mitochondrial outer membrane permeabilization (MOMP). Here we show that MTCH2/MIMP (mitochondrial carrier homologue 2/Met-induced mitochondrial protein), a novel truncated BID (tBID)-interacting protein, is a surface-exposed outer mitochondrial membrane protein that facilitates the recruitment of tBID to mitochondria. Knockout of MTCH2/MIMP in embryonic stem cells and in mouse embryonic fibroblasts hinders the recruitment of tBID to mitochondria, the activation of Bax/Bak, MOMP, and apoptosis. Moreover, conditional knockout of MTCH2/MIMP in the liver decreases the sensitivity of mice to Fas-induced hepatocellular apoptosis and prevents the recruitment of tBID to liver mitochondria both in vivo and in vitro. In contrast, MTCH2/MIMP deletion had no effect on apoptosis induced by other pro-apoptotic Bcl-2 family members and no detectable effect on the outer membrane lipid composition. These loss-of-function models indicate that MTCH2/MIMP has a critical function in liver apoptosis by regulating the recruitment of tBID to mitochondria.

To identify the structural features required for regulation of the mitochondrial permeability transition pore (PTP) by ubiquinone analogs (Fontaine, E., Ichas, F., and Bernardi, P. (1998) J. Biol. Chem. 40, 25734 -25740), we have carried out an analysis with quinone structural variants. We show that three functional classes can be defined: (i) PTP inhibitors (ubiquinone 0, decylubiquinone, ubiquinone 10, 2,3-dimethyl-6-decyl-1,4-benzoquinone, and 2,3,5-trimethyl-6-geranyl-1,4-benzoquinone); (ii) PTP inducers (2,3-dimethoxy-5-methyl-6-(10-hydroxydecyl)-1,4-benzoquinone and 2,5-dihydroxy-6-undecyl-1,4-benzoquinone); and (iii) PTP-inactive quinones that counteract the effects of both inhibitors and inducers (ubiquinone 5 and 2,3,5-trimethyl-6-(3-hydroxyisoamyl)-1,4-benzoquinone). The structure-function correlation indicates that minor modifications in the isoprenoid side chain can turn an inhibitor into an activator, and that the methoxy groups are not essential for the effects of quinones on the PTP. Since the ubiquinone analogs used in this study have a similar midpoint potential and decrease mitochondrial production of reactive oxygen species to the same extent, these results support the hypothesis that quinones modulate the PTP through a common binding site rather than through oxidation-reduction reactions. Occupancy of this site can modulate the PTP open-closed transitions, possibly through secondary changes of the PTP Ca 2؉ binding affinity.Regulation of ion fluxes across the inner mitochondrial membrane is essential both for metabolic regulation and for energy conservation. The inner membrane possesses an intrinsically low permeability to ions and solutes, which allows energy conservation in the form of a proton electrochemical potential difference (1), and a set of channels and transporters that regulate ion fluxes and volume homeostasis (see Ref. 2 for a recent review). However, mitochondria in vitro can easily undergo a permeability increase to solutes with molecular masses of about 1,500 Da or lower, which is followed by deenergization, disruption of ionic homeostasis, and swelling, the so-called permeability transition (PT).

The fundamental contribution of the mitochondria and ER to the decision made on the cell's fate has been increasingly recognized. This progress has illuminated the need for the mechanisms these organelles use to initiate and to propagate apoptotic signals. The toolbox of the mitochondria and ER is evolutionary conserved, overlapping and complementary. Furthermore, mitochondria are often closely associated with the ER providing the conditions for a local and privileged communication between the two organelles. The present review is concerned with the spatially and temporally coordinated utilization of Bcl-2 family proteins and Ca(2+) by the mitochondria and ER to control the membrane permeabilization in the mitochondria and to regulate Ca(2+) distribution and the activity of apoptotic proteins in the ER. The apoptotic means of the mitochondria and ER will eventually come together to control the dismantling of the cell by the caspases and other enzymes.

Recent findings indicate that perturbations of the mitochondrial electron transport chain (METC) can cause extended longevity in evolutionarily diverse organisms. To uncover the molecular basis of how altered METC increases lifespan in C. elegans, we performed an RNAi screen and revealed that three predicted transcription factors are specifically required for the extended longevity of mitochondrial mutants. In particular, we demonstrated that the nuclear homeobox protein CEH-23 uniquely mediates the longevity but not the slow development, reduced brood size, or resistance to oxidative stress associated with mitochondrial mutations. Furthermore, we showed that ceh-23 expression levels are responsive to altered METC, and enforced overexpression of ceh-23 is sufficient to extend lifespan in wild-type background. Our data point to mitochondria-to-nucleus communications to be key for longevity determination and highlight CEH-23 as a novel longevity factor capable of responding to mitochondrial perturbations. These findings provide a new paradigm for how mitochondria impact aging and age-dependent diseases.

SummaryHere, we report that inactivation of the Caenorhabditis elegans dynamin-related protein DRP-1, a key component responsible for mitochondrial fission and conserved from yeast to humans, dramatically enhanced the effect of reduced insulin signaling (IIS) to extend lifespan. This represents the first report of a beneficial impact of manipulating mitochondrial dynamics on animal lifespan and suggests that mitochondrial morphology and IIS cooperate to modulate aging. Key words: aging; Caenorhabditis elegans; fission protein DRP-1; insulin signaling; mitochondria.Mitochondria are dynamic organelles able to undergo frequent morphological and numeral changes. A delicate balance between mitochondrial fusion and fission is critical for broad aspects of animal physiology, including apoptosis and control of mitochondrial inheritance and quality (Seo et al., 2010). From yeast to humans, deregulations of mitochondrial network equilibrium as evident by disrupted mitochondrial morphology and accumulation of abnormally shaped mitochondria have been associated with senescence, aging, and aging-related diseases (Sohal, 1975;Yasuda et al., 2006;Lee et al., 2007). However, no direct evidence implicates mitochondrial dynamics in longevity determination in animals. The only evidence that mitochondrial plasticity positively impacts lifespan was shown in fungal models, in which reduced mitochondrial fission led to increased lifespan (Scheckhuber et al., 2007;Palermo et al., 2010). Mitochondrial dynamics are governed by molecular machineries that are highly conserved (Okamoto & Shaw, 2005). The dynaminrelated protein DRP-1 is the only profission protein identified in Caenorhabditis elegans and is demonstrated to control the scission of the mitochondrial outer membrane (Labrousse et al., 1999;Westermann, 2010).To explore how mitochondrial fission can impact animal lifespan, we monitored the lifespan of worms either treated with drp-1 RNAi or bearing a putative null mutation in the drp-1 gene (Breckenridge et al., 2008). The mean lifespan of wild-type worms treated with drp-1 RNAi or drp-1-mutant worms was indistinguishable from that of control animals ( Fig. 1A,B, Table S1, Supporting information), even though their mitochondrial morphology is greatly disrupted (Labrousse et al., 1999; Figs 2B,C and S1, Supporting information), indicating that reduced mitochondrial fission does not affect C. elegans lifespan under normal culturing condition.The insulin ⁄ IGF-1 signaling (IIS) pathway is a key longevity pathway, and C. elegans mutants with reduced IIS, such as the phosphatidyl-inositol 3-kinase age-1 mutant (Friedman & Johnson, 1988;Morris et al., 1996) and the tyrosine kinase insulin ⁄ IGF receptor daf-2 mutant (Kenyon et al., 1993), are longlived ( Fig. 1 A,C,D). Given that mitochondrial fission plays a critical role in insulin secretion in mammals (Yoon et al., 2011), we next tested how inactivating drp-1 might affect the longevity of IIS mutants. Strikingly, both age-1 and daf-2 mutants treated with drp-1 RNAi showed substant...

The permeability transition pore (PTP) is a mitochondrial inner membrane Ca2+-sensitive channel that plays a key role in different models of cell death. In a series of recent studies we have shown that the PTP is modulated by quinones, and we have identified three functional classes: (i) PTP inhibitors; (ii) PTP inducers; and (iii) PTP-inactive quinones that compete with both inhibitors and inducers. Here, we review our current understanding of pore regulation by quinones, and present the results obtained with a new series of structural variants. Based on the effects of the compounds studied so far, we confirm that minor structural changes profoundly modify the effects of quinones on the PTP. On the other hand, quinones with very different structural features may have qualitatively similar effects on the PTP. Taken together, these results support our original proposal that quinones affect the PTP through a common binding site whose occupancy modulates its open-closed transitions, possibly through secondary changes of the Ca2+-binding affinity.