Docosahexaenoic acid lowers cardiac mitochondrial enzyme activity by replacing linoleic acid in the phospholipidome

Cardiac mitochondrial phospholipid acyl chains regulate respiratory enzymatic activity. In several diseases, the rodent cardiac phospholipidome is extensively rearranged; however, whether specific acyl chains impair respiratory enzyme function is unknown. One unique remodeling event in the myocardium of obese and diabetic rodents is an increase in docosahexaenoic acid (DHA) levels. Here, we first confirmed that cardiac DHA levels are elevated in diabetic humans relative to controls. We then used dietary supplementation of a Western diet with DHA as a tool to promote cardiac acyl chain remodeling and to study its influence on respiratory enzyme function. DHA extensively remodeled the acyl chains of cardiolipin (CL), mono-lyso CL, phosphatidylcholine, and phosphatidylethanolamine. Moreover, DHA lowered enzyme activities of respiratory complexes I, IV, V, and I+III. Mechanistically, the reduction in enzymatic activities were not driven by a dramatic reduction in the abundance of supercomplexes. Instead, replacement of tetralinoleoyl-CL with tetradocosahexaenoyl-CL in biomimetic membranes prevented formation of phospholipid domains that regulate enzyme activity. Tetradocosahexaenoyl-CL inhibited domain organization due to favorable Gibbs free energy of phospholipid mixing. Furthermore, in vitro substitution of tetralinoleoyl-CL with tetradocosahexaenoyl-CL blocked complex-IV binding. Finally, reintroduction of linoleic acid, via fusion of phospholipid vesicles to mitochondria isolated from DHA-fed mice, rescued the major losses in the mitochondrial phospholipidome and complexes I, IV, and V activities. Altogether, our results show that replacing linoleic acid with DHA lowers select cardiac enzyme activities by potentially targeting domain organization and phospholipid-protein binding, which has implications for the ongoing debate about polyunsaturated fatty acids and cardiac health

The Mitochondrial Ca(2+) Uniporter: Structure, Function, and Pharmacology.

Mitochondrial Ca(2+) uptake is crucial for an array of cellular functions while an imbalance can elicit cell death. In this chapter, we briefly reviewed the various modes of mitochondrial Ca(2+) uptake and our current understanding of mitochondrial Ca(2+) homeostasis in regards to cell physiology and pathophysiology. Further, this chapter focuses on the molecular identities, intracellular regulators as well as the pharmacology of mitochondrial Ca(2+) uniporter complex

Basic Atomic Physics

Contains reports on seven research projects.National Science Foundation (Grant PHY 87-06560)Joint Services Electronics Program (Contract DAAL03-86-K-0001)Joint Services Electronics Program (Contract DAAL03-89-C-0002)National Science Foundation (Grant PHY 86-05893)U.S. Navy - Office of Naval Research (Contract N00014-83-K-0695)U.S. Navy - Office of Naval Research (Contract N00014-89-J-1207

MICU2, a Paralog of MICU1, Resides within the Mitochondrial Uniporter Complex to Regulate Calcium Handling

Mitochondrial calcium uptake is present in nearly all vertebrate tissues and is believed to be critical in shaping calcium signaling, regulating ATP synthesis and controlling cell death. Calcium uptake occurs through a channel called the uniporter that resides in the inner mitochondrial membrane. Recently, we used comparative genomics to identify MICU1 and MCU as the key regulatory and putative pore-forming subunits of this channel, respectively. Using bioinformatics, we now report that the human genome encodes two additional paralogs of MICU1, which we call MICU2 and MICU3, each of which likely arose by gene duplication and exhibits distinct patterns of organ expression. We demonstrate that MICU1 and MICU2 are expressed in HeLa and HEK293T cells, and provide multiple lines of biochemical evidence that MCU, MICU1 and MICU2 reside within a complex and cross-stabilize each other's protein expression in a cell-type dependent manner. Using in vivo RNAi technology to silence MICU1, MICU2 or both proteins in mouse liver, we observe an additive impairment in calcium handling without adversely impacting mitochondrial respiration or membrane potential. The results identify MICU2 as a new component of the uniporter complex that may contribute to the tissue-specific regulation of this channel.National Institutes of Health (U.S.) (GM0077465)National Institutes of Health (U.S.) (DK080261

Improved Mitochondrial Function with Diet-Induced Increase in Either Docosahexaenoic Acid or Arachidonic Acid in Membrane Phospholipids

Mitochondria can depolarize and trigger cell death through the opening of the mitochondrial permeability transition pore (MPTP). We recently showed that an increase in the long chain n3 polyunsaturated fatty acids (PUFA) docosahexaenoic acid (DHA; 22:6n3) and depletion of the n6 PUFA arachidonic acid (ARA; 20:4n6) in mitochondrial membranes is associated with a greater Ca2+ load required to induce MPTP opening. Here we manipulated mitochondrial phospholipid composition by supplementing the diet with DHA, ARA or combined DHA+ARA in rats for 10 weeks. There were no effects on cardiac function, or respiration of isolated mitochondria. Analysis of mitochondrial phospholipids showed DHA supplementation increased DHA and displaced ARA in mitochondrial membranes, while supplementation with ARA or DHA+ARA increased ARA and depleted linoleic acid (18:2n6). Phospholipid analysis revealed a similar pattern, particularly in cardiolipin. Tetralinoleoyl cardiolipin was depleted by 80% with ARA or DHA+ARA supplementation, with linoleic acid side chains replaced by ARA. Both the DHA and ARA groups had delayed Ca2+-induced MPTP opening, but the DHA+ARA group was similar to the control diet. In conclusion, alterations in mitochondria membrane phospholipid fatty acid composition caused by dietary DHA or ARA was associated with a greater cumulative Ca2+ load required to induced MPTP opening. Further, high levels of tetralinoleoyl cardiolipin were not essential for normal mitochondrial function if replaced with very-long chain n3 or n6 PUFAs

The Effect of OPA1 on Mitochondrial Ca2+ Signaling

The dynamin-related GTPase protein OPA1, localized in the intermembrane space and tethered to the inner membrane of mitochondria, participates in the fusion of these organelles. Its mutation is the most prevalent cause of Autosomal Dominant Optic Atrophy. OPA1 controls the diameter of the junctions between the boundary part of the inner membrane and the membrane of cristae and reduces the diffusibility of cytochrome c through these junctions. We postulated that if significant Ca2+ uptake into the matrix occurs from the lumen of the cristae, reduced expression of OPA1 would increase the access of Ca2+ to the transporters in the crista membrane and thus would enhance Ca2+ uptake. In intact H295R adrenocortical and HeLa cells cytosolic Ca2+ signals evoked with K+ and histamine, respectively, were transferred into the mitochondria. The rate and amplitude of mitochondrial [Ca2+] rise (followed with confocal laser scanning microscopy and FRET measurements with fluorescent wide-field microscopy) were increased after knockdown of OPA1, as compared with cells transfected with control RNA or mitofusin1 siRNA. Ca2+ uptake was enhanced despite reduced mitochondrial membrane potential. In permeabilized cells the rate of Ca2+ uptake by depolarized mitochondria was also increased in OPA1-silenced cells. The participation of Na+/Ca2+ and Ca2+/H+ antiporters in this transport process is indicated by pharmacological data. Altogether, our observations reveal the significance of OPA1 in the control of mitochondrial Ca2+ metabolism

Proteomic profiling of cellular steatosis with concomitant oxidative stress in vitro

10.1186/s12944-016-0283-7Lipids in Health and Disease15111