Oxidant stress induces constitutive calcium entry by tacking glutathiones onto the Orai CRAC channel activator STIM1.

SUMMARY Ca2+ dynamics and oxidative signaling are fundamental mechanisms for mitochondrial bioenergetics and cell function. The MCU complex is the major pathway by which these signals are integrated in mitochondria. Whether and how these coactive elements interact with MCU has not been established. As an approach towards understanding the regulation of MCU channel by oxidative milieu, we adapted inflammatory and hypoxia models. We identified the conserved cysteine 97 to be the only reactive thiol in human MCU that undergoes S-glutathionylation. Furthermore, biochemical, structural and superresolution imaging analysis revealed that MCU oxidation promotes MCU higher-order oligomer formation. Both oxidation and mutation of MCU Cys-97 exhibited persistent MCU channel activity with higher [Ca2+]m uptake rate, elevated mROS and enhanced [Ca2+]m overload-induced cell death. In contrast, these effects were largely independent of MCU interaction with its regulators. These findings reveal a distinct functional role for Cys-97 in ROS sensing and regulation of MCU activity.

SUMMARY Mitochondria shape cytosolic calcium ([Ca2+]c) transients and utilize the mitochondrial Ca2+ ([Ca2+]m) in exchange for bioenergetics output. Conversely, dys-regulated [Ca2+]c causes [Ca2+]m overload and induces permeability transition pore and cell death. Ablation of MCU-mediated Ca2+ uptake exhibited elevated [Ca2+]c and failed to prevent stress-induced cell death. The mechanisms for these effects remain elusive. Here, we report that mitochondria undergo a cytosolic Ca2+-induced shape change that is distinct from mitochondrial fission and swelling. [Ca2+]c elevation, but not MCU-mediated Ca2+ uptake, appears to be essential for the process we term mitochondrial shape transition (MiST). MiST is mediated by the mitochondrial protein Miro1 through its EF-hand domain 1 in multiple cell types. Moreover, Ca2+-dependent disruption of Miro1/KIF5B/tubulin complex is determined by Miro1 EF1 domain. Functionally, Miro1-dependent MiST is essential for autophagy/mitophagy that is attenuated in Miro1 EF1 mutants. Thus, Miro1 is a cytosolic Ca2+ sensor that decodes metazoan Ca2+ signals as MiST.

Rationale: Peroxisome proliferator-activated receptors (PPARs) (␣, ␥, and ␦/␤) are nuclear hormone receptors and ligand-activated transcription factors that serve as key determinants of myocardial fatty acid metabolism. Long-term cardiomyocyte-restricted PPAR␦ deficiency in mice leads to depressed myocardial fatty acid oxidation, bioenergetics, and premature death with lipotoxic cardiomyopathy. Objective: To explore the essential role of PPAR␦ in the adult heart. Methods and Results: We investigated the consequences of inducible short-term PPAR␦ knockout in the adult mouse heart. In addition to a substantial transcriptional downregulation of lipid metabolic proteins, short-term PPAR␦ knockout in the adult mouse heart attenuated cardiac expression of both Cu/Zn superoxide dismutase and manganese superoxide dismutase, leading to increased oxidative damage to the heart. Moreover, expression of key mitochondrial biogenesis determinants such as PPAR␥ coactivator-1 were substantially decreased in the short-term PPAR␦ deficient heart, concomitant with a decreased mitochondrial DNA copy number. Rates of palmitate and glucose oxidation were markedly depressed in cardiomyocytes of PPAR␦ knockout hearts. Consequently, PPAR␦ deficiency in the adult heart led to depressed cardiac performance and cardiac hypertrophy. Conclusions: PPAR␦ is an essential regulator of cardiac mitochondrial protection and biogenesis and PPAR␦ activation can be a potential therapeutic target for cardiac disorders. (Circ Res. 2010;106:911-919.)

Nitric oxide and other reactive nitrogen species target multiple sites in the mitochondria to impact cellular bioenergetics and survival. Kinetic imaging studies revealed that NO from either activated macrophages or donor compounds rapidly diffuses to the mitochondria, causing a dose dependent progressive increase in NO-dependent DAF fluorescence that corresponded to mitochondrial membrane potential loss, and initiated alterations in cellular bioenergetics that ultimately led to necrotic cell death. Cellular dysfunction is mediated by an elevated 3-nitrotyrosine signature of the mitochondrial complex I subunit NDUFB8, which is vital for normal mitochondrial function as evidenced by selective knockdown via siRNA. Overexpression of mitochondrial superoxide dismutase substantially decreased NDUFB8 nitration and restored mitochondrial homeostasis. Further, treatment of cells with either necrostatin-1 or siRNA knockdown of RIP1 and RIP3 prevented NO-mediated necrosis. This work demonstrates that the interaction between NO and mitochondrially-derived superoxide alters mitochondrial bioenergetics and cell function, thus providing a molecular mechanism for reactive oxygen and nitrogen species-mediated alterations in mitochondrial homeostasis.

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BackgroundOrganophosphates are the most frequently and largely applied insecticide in the world due to their biodegradable nature. Gut microbes were shown to degrade organophosphates and cause intestinal dysfunction. The diabetogenic nature of organophosphates was recently reported but the underlying molecular mechanism is unclear. We aimed to understand the role of gut microbiota in organophosphate-induced hyperglycemia and to unravel the molecular mechanism behind this process.ResultsHere we demonstrate a high prevalence of diabetes among people directly exposed to organophosphates in rural India (n = 3080). Correlation and linear regression analysis reveal a strong association between plasma organophosphate residues and HbA1c but no association with acetylcholine esterase was noticed. Chronic treatment of mice with organophosphate for 180 days confirms the induction of glucose intolerance with no significant change in acetylcholine esterase. Further fecal transplantation and culture transplantation experiments confirm the involvement of gut microbiota in organophosphate-induced glucose intolerance. Intestinal metatranscriptomic and host metabolomic analyses reveal that gut microbial organophosphate degradation produces short chain fatty acids like acetic acid, which induces gluconeogenesis and thereby accounts for glucose intolerance. Plasma organophosphate residues are positively correlated with fecal esterase activity and acetate level of human diabetes.ConclusionCollectively, our results implicate gluconeogenesis as the key mechanism behind organophosphate-induced hyperglycemia, mediated by the organophosphate-degrading potential of gut microbiota. This study reveals the gut microbiome-mediated diabetogenic nature of organophosphates and hence that the usage of these insecticides should be reconsidered.Electronic supplementary materialThe online version of this article (doi:10.1186/s13059-016-1134-6) contains supplementary material, which is available to authorized users.

Ethanol intoxication stimulates the production of proinflammatory cytokines, increases the formation of reactive oxygen species, and induces mitochondrial impairment. However, information is limited as to the exact sequence and components involved in ethanol-induced hepatotoxicity. Acute ethanol exposure enhances mitochondrial superoxide (O2 .−) production and impairs mitochondrial Ca2+ handling. In turn, O2 .− facilitates cytochrome c release and mitochondrial membrane potential loss that is not dependent upon H2O2 and divalent cations and requires Bak in a Bax-independent fashion. Furthermore, triggering of Bak's proapoptotic activity requires the cytosolic presence of Bid, a BH3-only protein that is processed by the initiator caspase-2. Together, these studies identify an O2 .−-driven, caspase-initiated apoptotic pathway that selectively involves the Bcl-2 family proteins Bid and Bak. This pathway manifests itself during chronic ethanol consumption in aged animals and identifies caspase-2, Bid, and Bak as essential mediators of O2 .−-induced apoptosis that may prove effective targets for the development of therapeutics to treat alcoholic liver disease.