MCU-complex-mediated mitochondrial calcium signaling is impaired in Barth syndrome

Abstract Calcium signaling via mitochondrial calcium uniporter (MCU) complex coordinates mitochondrial bioenergetics with cellular energy demands. Emerging studies show that the stability and activity of the pore-forming subunit of the complex, MCU, is dependent on the mitochondrial phospholipid, cardiolipin (CL), but how this impacts calcium-dependent mitochondrial bioenergetics in CL-deficiency disorder like Barth syndrome (BTHS) is not known. Here we utilized multiple models of BTHS including yeast, mouse muscle cell line, as well as BTHS patient cells and cardiac tissue to show that CL is required for the abundance and stability of the MCU-complex regulatory subunit MICU1. Interestingly, the reduction in MICU1 abundance in BTHS mitochondria is independent of MCU. Unlike MCU and MICU1/MICU2, other subunit and associated factor of the uniporter complex, EMRE and MCUR1, respectively, are not affected in BTHS models. Consistent with the decrease in MICU1 levels, we show that the kinetics of MICU1-dependent mitochondrial calcium uptake is perturbed and acute stimulation of mitochondrial calcium signaling in BTHS myoblasts fails to activate pyruvate dehydrogenase, which in turn impairs the generation of reducing equivalents and blunts mitochondrial bioenergetics. Taken together, our findings suggest that defects in mitochondrial calcium signaling could contribute to cardiac and skeletal muscle pathologies observed in BTHS patients.

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Abstract P467: Cardiac-specific Deletion Of Voltage Dependent Anion Channel 2 Leads To Dilated Cardiomyopathy By Altering Calcium Homeostasis

Circulation Research ◽

10.1161/res.129.suppl_1.p467 ◽

2021 ◽

Vol 129 (Suppl_1) ◽

Author(s):

Thirupura S Shankar ◽

Dinesh Kumar Anandamurugan Ramadurai ◽

Kira Steinhorst ◽

Salah Sommakia ◽

Rachit Badolia ◽

...

Keyword(s):

Heart Failure ◽

Dilated Cardiomyopathy ◽

Calcium Signaling ◽

Calcium Homeostasis ◽

Calcium Uptake ◽

Mitochondrial Calcium ◽

Voltage Dependent Anion Channel ◽

Knock Out ◽

Cellular Calcium ◽

Mitochondrial Calcium Uptake

Voltage dependent anion channel 2 (VDAC2) is a mitochondrial outer membrane porin known to play a significant role in apoptosis and calcium signaling. Abnormalities in cellular calcium homeostasis often leads to electrical and contractile dysfunction and can cause dilated cardiomyopathy and heart failure. Previous literature suggests that improving mitochondrial calcium uptake via VDAC2 rescues arrhythmia phenotypes in genetic models of impaired cellular calcium signaling. However, the direct role of VDAC2 in intracellular calcium signaling and cardiac function is not well understood. To elucidate the role of VDAC2 in calcium homeostasis, we generated a cardiac-specific deletion of Vdac2 in mice. Our results indicate that loss of VDAC2 in the myocardium during development causes severe impairment in excitation-contraction coupling by reducing mitochondrial calcium uptake (n=3, p<0.05) and thereby impairing intracellular calcium signaling. VDAC2 knock-out mice showed a significant reduction in RYR-mediated calcium release (F/F 0 ) and rate of calcium uptake by SERCA2a [tau(msec)] compared to control mice (N=3, WT=54, KO=38, p<0.0001 (F/F 0 ) and p<0.05 (tau)). We also observed adverse cardiac remodeling which progressed to severe dilated cardiomyopathy and death (N=6, p<0.0001). Reintroducing VDAC2 in 6-week-old knock-out mice partially rescued the cardiomyopathy phenotype evident from improvement in ejection fraction and fractional shortening (n=3, p<0.05). Improving mitochondrial calcium uptake via VDAC2 using a VDAC2 agonist efsevin, increased cardiac contractile force in a mouse model of pressure-overload induced heart failure (N=8, n=22, p<0.05). In conclusion, our findings demonstrate that VDAC2 plays a crucial role in cardiac function by influencing mitochondrial and cellular calcium signaling. Through this role in cellular calcium dynamics and excitation-contraction coupling VDAC2 emerges as a plausible therapeutic target for heart failure.

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Abstract P019: Renal Mitochondrial Bioenergetics In Salt-sensitive Hypertension Is Affected By ANP

Hypertension ◽

10.1161/hyp.76.suppl_1.p019 ◽

2020 ◽

Vol 76 (Suppl_1) ◽

Author(s):

Regina Sultanova ◽

Mark Domondon ◽

Anna Nikiforova ◽

Ryan Schibalski ◽

Daria Ilatovskaya

Keyword(s):

Mitochondrial Function ◽

Calcium Influx ◽

Kidney Injury ◽

Mitochondrial Calcium ◽

Mitochondrial Bioenergetics ◽

Amplex Red ◽

Respiratory Chain Activity ◽

Mitochondrial Calcium Uptake ◽

Salt Sensitive Hypertension

There are clinical data suggesting that low levels of Atrial Natriuretic Peptide (ANP) aggravate susceptibility to salt-sensitive (SS) hypertension. ANP is known to affect mitochondria in many tissues, however, little is known about the effects of ANP on renal mitochondrial function. According to our earlier studies, Dahl SS rats lacking ANP exhibit increased blood pressure and pronounced kidney injury. We hypothesized that in SS hypertension ANP deficiency affects renal mitochondrial bioenergetics and contributes to renal function impairment. SS hypertension was induced in male SS NPPA-/- ( Nppa knockout in Dahl SS background, KO) and SS WT (wild type, WT) rats by a high salt (HS) 4% NaCl diet administered for 21 days. Age-matched control animals were fed a normal (NS) 0.4% NaCl salt diet. A combination of in vivo studies, molecular biology and tests of mitochondria isolated from renal cortex (seahorse respiration and spectrofluorimetry assays using TMRM, Amplex Red and MCLA) were used to probe the role ANP in mitochondrial function. Data was analyzed with ANOVA followed by Holm-Sidak post hoc. We report a significant decrease in mitochondrial membrane potential in the SS NPPA-/- rats vs SS WT (25 ± 4% decrease in KO vs WT on NS, and a 16 ± 5% decrease in KO on HS). Furthermore, mitochondrial H 2 O 2 (57.7 ± 7.4 (WT) vs 57.6 ± 1.3 au (KO), p<0.0001) and superoxide (36.6± 1.8 au (WT) vs 67.9 ± 5.2 au (KO), p<0.0001) levels were increased in the KO on a HS diet. Next, we antioxidant capacity was elevated in the SS NPPA-/- rats on HS diet in comparison with SS WT , and increased SOD2 levels were observed in the KO animals. Furthermore, SS NPPA-/- rats exhibit higher MCU (mitochondrial calcium uniporter) activity than SS WT , implying an ANP-dependent effect on mitochondrial calcium influx. Seahorse assay revealed dramatic elevation of the basal, ATP-linked, maximal and spare oxygen consumption rate (OCR) in the cortical mitochondria of the knockout rats on HS, while in HS fed SS WT rats these OCR parameters were, as expected, reduced (vs NS). Therefore, deficiency of circulating ANP leads to significant changes of renal mitochondrial bioenergetics, potentially via effects on mitochondrial calcium uptake, which in turn affects respiratory chain activity leading to oxidative stress.

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ER–Mitochondria contact sites: A new regulator of cellular calcium flux comes into play

The Journal of Cell Biology ◽

10.1083/jcb.201607124 ◽

2016 ◽

Vol 214 (4) ◽

pp. 367-370 ◽

Cited By ~ 50

Author(s):

Michiel Krols ◽

Geert Bultynck ◽

Sophie Janssens

Keyword(s):

Endoplasmic Reticulum ◽

Calcium Signaling ◽

Calcium Pump ◽

Calcium Flux ◽

Mitochondrial Calcium ◽

Mitochondrial Bioenergetics ◽

Cellular Calcium ◽

Contact Sites ◽

Cell Biol ◽

Membrane Contacts

Endoplasmic reticulum (ER)–mitochondria membrane contacts are hotspots for calcium signaling. In this issue, Raturi et al. (2016. J. Cell Biol. http://dx.doi.org/10.1083/jcb.201512077) show that the thioredoxin TMX1 inhibits the calcium pump SERCA2b at ER–mitochondria contact sites, thereby affecting ER–mitochondrial calcium transfer and mitochondrial bioenergetics.

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An essential role for cardiolipin in the stability and function of the mitochondrial calcium uniporter

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2000640117 ◽

2020 ◽

Vol 117 (28) ◽

pp. 16383-16390 ◽

Cited By ~ 9

Author(s):

Sagnika Ghosh ◽

Writoban Basu Ball ◽

Travis R. Madaris ◽

Subramanya Srikantan ◽

Muniswamy Madesh ◽

...

Keyword(s):

Calcium Transport ◽

Cardiac Tissue ◽

Barth Syndrome ◽

Mitochondrial Calcium ◽

Mitochondrial Calcium Uniporter ◽

Cellular Models ◽

Calcium Uniporter ◽

Protein Components ◽

And Function

Calcium uptake by the mitochondrial calcium uniporter coordinates cytosolic signaling events with mitochondrial bioenergetics. During the past decade all protein components of the mitochondrial calcium uniporter have been identified, including MCU, the pore-forming subunit. However, the specific lipid requirements, if any, for the function and formation of this channel complex are currently not known. Here we utilize yeast, which lacks the mitochondrial calcium uniporter, as a model system to address this problem. We use heterologous expression to functionally reconstitute human uniporter machinery both in wild-type yeast as well as in mutants defective in the biosynthesis of phosphatidylethanolamine, phosphatidylcholine, or cardiolipin (CL). We uncover a specific requirement of CL for in vivo reconstituted MCU stability and activity. The CL requirement of MCU is evolutionarily conserved with loss of CL triggering rapid turnover of MCU homologs and impaired calcium transport. Furthermore, we observe reduced abundance and activity of endogenous MCU in mammalian cellular models of Barth syndrome, which is characterized by a partial loss of CL. MCU abundance is also decreased in the cardiac tissue of Barth syndrome patients. Our work raises the hypothesis that impaired mitochondrial calcium transport contributes to the pathogenesis of Barth syndrome, and more generally, showcases the utility of yeast phospholipid mutants in dissecting the phospholipid requirements of ion channel complexes.

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