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2.
Sci Rep ; 14(1): 11497, 2024 05 20.
Article in English | MEDLINE | ID: mdl-38769106

ABSTRACT

Barth syndrome (BTHS) is a rare disorder caused by mutations in the TAFAZZIN gene. Previous studies from both patients and model systems have established metabolic dysregulation as a core component of BTHS pathology. In particular, features such as lactic acidosis, pyruvate dehydrogenase (PDH) deficiency, and aberrant fatty acid and glucose oxidation have been identified. However, the lack of a mechanistic understanding of what causes these conditions in the context of BTHS remains a significant knowledge gap, and this has hindered the development of effective therapeutic strategies for treating the associated metabolic problems. In the current study, we utilized tafazzin-knockout C2C12 mouse myoblasts (TAZ-KO) and cardiac and skeletal muscle tissue from tafazzin-knockout mice to identify an upstream mechanism underlying impaired PDH activity in BTHS. This mechanism centers around robust upregulation of pyruvate dehydrogenase kinase 4 (PDK4), resulting from hyperactivation of AMP-activated protein kinase (AMPK) and subsequent transcriptional upregulation by forkhead box protein O1 (FOXO1). Upregulation of PDK4 in tafazzin-deficient cells causes direct phospho-inhibition of PDH activity accompanied by increased glucose uptake and elevated intracellular glucose concentration. Collectively, our findings provide a novel mechanistic framework whereby impaired tafazzin function ultimately results in robust PDK4 upregulation, leading to impaired PDH activity and likely linked to dysregulated metabolic substrate utilization. This mechanism may underlie previously reported findings of BTHS-associated metabolic dysregulation.


Subject(s)
AMP-Activated Protein Kinases , Forkhead Box Protein O1 , Mice, Knockout , Pyruvate Dehydrogenase Acetyl-Transferring Kinase , Animals , Mice , Forkhead Box Protein O1/metabolism , Forkhead Box Protein O1/genetics , AMP-Activated Protein Kinases/metabolism , Pyruvate Dehydrogenase Acetyl-Transferring Kinase/metabolism , Pyruvate Dehydrogenase Acetyl-Transferring Kinase/genetics , Transcription Factors/metabolism , Transcription Factors/genetics , Up-Regulation , Signal Transduction , Myoblasts/metabolism , Cell Line , Glucose/metabolism , Acyltransferases
3.
bioRxiv ; 2024 Feb 04.
Article in English | MEDLINE | ID: mdl-38352304

ABSTRACT

Barth syndrome (BTHS) is a rare disorder caused by mutations in the TAFAZZIN gene. Previous studies from both patients and model systems have established metabolic dysregulation as a core component of BTHS pathology. In particular, features such as lactic acidosis, pyruvate dehydrogenase (PDH) deficiency, and aberrant fatty acid and glucose oxidation have been identified. However, the lack of a mechanistic understanding of what causes these conditions in the context of BTHS remains a significant knowledge gap, and this has hindered the development of effective therapeutic strategies for treating the associated metabolic problems. In the current study, we utilized tafazzin-knockout C2C12 mouse myoblasts (TAZ-KO) and cardiac and skeletal muscle tissue from tafazzin-knockout mice to identify an upstream mechanism underlying impaired PDH activity in BTHS. This mechanism centers around robust upregulation of pyruvate dehydrogenase kinase 4 (PDK4), resulting from hyperactivation of AMP-activated protein kinase (AMPK) and subsequent transcriptional upregulation by forkhead box protein O1 (FOXO1). Upregulation of PDK4 in tafazzin-deficient cells causes direct phospho-inhibition of PDH activity accompanied by increased glucose uptake and elevated intracellular glucose concentration. Collectively, our findings provide a novel mechanistic framework whereby impaired tafazzin function ultimately results in robust PDK4 upregulation, leading to impaired PDH activity and likely linked to dysregulated metabolic substrate utilization. This mechanism may underlie previously reported findings of BTHS-associated metabolic dysregulation.

4.
J Biol Chem ; 299(10): 105241, 2023 10.
Article in English | MEDLINE | ID: mdl-37690688

ABSTRACT

Respiratory complexes and cardiolipins have exceptionally long lifetimes. The fact that they co-localize in mitochondrial cristae raises the question of whether their longevities have a common cause and whether the longevity of OXPHOS proteins is dependent on cardiolipin. To address these questions, we developed a method to measure side-by-side the half-lives of proteins and lipids in wild-type Drosophila and cardiolipin-deficient mutants. We fed adult flies with stable isotope-labeled precursors (13C615N2-lysine or 13C6-glucose) and determined the relative abundance of heavy isotopomers in protein and lipid species by mass spectrometry. To minimize the confounding effects of tissue regeneration, we restricted our analysis to the thorax, the bulk of which consists of post-mitotic flight muscles. Analysis of 680 protein and 45 lipid species showed that the subunits of respiratory complexes I-V and the carriers for phosphate and ADP/ATP were among the longest-lived proteins (average half-life of 48 ± 16 days) while the molecular species of cardiolipin were the longest-lived lipids (average half-life of 27 ± 6 days). The remarkable longevity of these crista residents was not shared by all mitochondrial proteins, especially not by those residing in the matrix and the inner boundary membrane. Ablation of cardiolipin synthase, which causes replacement of cardiolipin by phosphatidylglycerol, and ablation of tafazzin, which causes partial replacement of cardiolipin by monolyso-cardiolipin, decreased the lifetimes of the respiratory complexes. Ablation of tafazzin also decreased the lifetimes of the remaining cardiolipin species. These data suggest that an important function of cardiolipin in mitochondria is to protect respiratory complexes from degradation.


Subject(s)
Cardiolipins , Animals , Cardiolipins/metabolism , Mitochondria/metabolism , Mitochondrial Membranes/metabolism , Muscles/metabolism , Drosophila melanogaster
5.
Cell Rep ; 42(8): 112846, 2023 08 29.
Article in English | MEDLINE | ID: mdl-37516961

ABSTRACT

Several phospholipid (PL) molecules are intertwined with some mitochondrial complex I (CI) subunits in the membrane domain of CI, but their function is unclear. We report that when the Drosophila melanogaster ortholog of the intramitochondrial PL transporter, STARD7, is severely disrupted, assembly of the oxidative phosphorylation (OXPHOS) system is impaired, and the biogenesis of several CI subcomplexes is hampered. However, intriguingly, a restrained knockdown of STARD7 impairs the incorporation of NDUFS5 and NDUFA1 into the proximal part of the CI membrane domain without directly affecting the incorporation of subunits in the distal part of the membrane domain, OXPHOS complexes already assembled, or mitochondrial cristae integrity. Importantly, the restrained knockdown of STARD7 appears to induce a modest amount of cardiolipin remodeling, indicating that there could be some alteration in the composition of the mitochondrial phospholipidome. We conclude that PLs can regulate CI biogenesis independent of their role in maintaining mitochondrial membrane integrity.


Subject(s)
Mitochondrial Membranes , Phospholipids , Animals , Mitochondrial Membranes/metabolism , Phospholipids/metabolism , Drosophila melanogaster/metabolism , Mitochondria/metabolism , Cardiolipins/metabolism , Oxidative Phosphorylation
6.
bioRxiv ; 2023 Mar 10.
Article in English | MEDLINE | ID: mdl-36945411

ABSTRACT

Background: Cardiomyocyte maturation requires a massive increase in respiratory enzymes and their assembly into long-lived complexes of oxidative phosphorylation (OXPHOS). The molecular mechanisms underlying the maturation of cardiac mitochondria have not been established. Methods: To determine whether the mitochondria-specific lipid cardiolipin is involved in cardiac maturation, we created a cardiomyocyte-restricted knockout (KO) of cardiolipin synthase ( Crls1 ) in mice and studied the postnatal development of the heart. We also measured the turnover rates of proteins and lipids in cardiolipin-deficient flight muscle from Drosophila, a tissue that has mitochondria with high OXPHOS activity like the heart. Results: Crls1KO mice survived the prenatal period but failed to accumulate OXPHOS proteins during postnatal maturation and succumbed to heart failure at the age of 2 weeks. Turnover measurements showed that the exceptionally long half-life of OXPHOS proteins is critically dependent on cardiolipin. Conclusions: Cardiolipin is essential for the postnatal maturation of cardiomyocytes because it allows mitochondrial cristae to accumulate OXPHOS proteins to a high concentration and to shield them from degradation.

7.
Hum Mol Genet ; 32(12): 2055-2067, 2023 06 05.
Article in English | MEDLINE | ID: mdl-36917259

ABSTRACT

Barth syndrome is an X-linked disorder caused by loss-of-function mutations in Tafazzin (TAZ), an acyltransferase that catalyzes remodeling of cardiolipin, a signature phospholipid of the inner mitochondrial membrane. Patients develop cardiac and skeletal muscle weakness, growth delay and neutropenia, although phenotypic expression varies considerably between patients. Taz knockout mice recapitulate many of the hallmark features of the disease. We used mouse genetics to test the hypothesis that genetic modifiers alter the phenotypic manifestations of Taz inactivation. We crossed TazKO/X females in the C57BL6/J inbred strain to males from eight inbred strains and evaluated the phenotypes of first-generation (F1) TazKO/Y progeny, compared to TazWT/Y littermates. We observed that genetic background strongly impacted phenotypic expression. C57BL6/J and CAST/EiJ[F1] TazKO/Y mice developed severe cardiomyopathy, whereas A/J[F1] TazKO/Y mice had normal heart function. C57BL6/J and WSB/EiJ[F1] TazKO/Y mice had severely reduced treadmill endurance, whereas endurance was normal in A/J[F1] and CAST/EiJ[F1] TazKO/Y mice. In all genetic backgrounds, cardiolipin showed similar abnormalities in knockout mice, and transcriptomic and metabolomic investigations identified signatures of mitochondrial uncoupling and activation of the integrated stress response. TazKO/Y cardiac mitochondria were small, clustered and had reduced cristae density in knockouts in severely affected genetic backgrounds but were relatively preserved in the permissive A/J[F1] strain. Gene expression and mitophagy measurements were consistent with reduced mitophagy in knockout mice in genetic backgrounds intolerant of Taz mutation. Our data demonstrate that genetic modifiers powerfully modulate phenotypic expression of Taz loss-of-function and act downstream of cardiolipin, possibly by altering mitochondrial quality control.


Subject(s)
Barth Syndrome , Male , Female , Animals , Mice , Barth Syndrome/genetics , Barth Syndrome/metabolism , Cardiolipins/metabolism , Transcription Factors/metabolism , Disease Models, Animal , Acyltransferases/genetics , Mice, Knockout , Phenotype
8.
J Biol Chem ; 299(3): 102978, 2023 03.
Article in English | MEDLINE | ID: mdl-36739949

ABSTRACT

The mitochondrial phospholipid cardiolipin (CL) is critical for numerous essential biological processes, including mitochondrial dynamics and energy metabolism. Mutations in the CL remodeling enzyme TAFAZZIN cause Barth syndrome, a life-threatening genetic disorder that results in severe physiological defects, including cardiomyopathy, skeletal myopathy, and neutropenia. To study the molecular mechanisms whereby CL deficiency leads to skeletal myopathy, we carried out transcriptomic analysis of the TAFAZZIN-knockout (TAZ-KO) mouse myoblast C2C12 cell line. Our data indicated that cardiac and muscle development pathways are highly decreased in TAZ-KO cells, consistent with a previous report of defective myogenesis in this cell line. Interestingly, the muscle transcription factor myoblast determination protein 1 (MyoD1) is significantly repressed in TAZ-KO cells and TAZ-KO mouse hearts. Exogenous expression of MyoD1 rescued the myogenesis defects previously observed in TAZ-KO cells. Our data suggest that MyoD1 repression is caused by upregulation of the MyoD1 negative regulator, homeobox protein Mohawk, and decreased Wnt signaling. Our findings reveal, for the first time, that CL metabolism regulates muscle differentiation through MyoD1 and identify the mechanism whereby MyoD1 is repressed in CL-deficient cells.


Subject(s)
Barth Syndrome , Cardiolipins , MyoD Protein , Animals , Mice , Acyltransferases/genetics , Barth Syndrome/genetics , Barth Syndrome/metabolism , Cardiolipins/genetics , Cardiolipins/metabolism , Mice, Knockout , Muscles/metabolism , Transcription Factors/metabolism , MyoD Protein/genetics , MyoD Protein/metabolism
9.
Dev Dyn ; 252(6): 691-712, 2023 06.
Article in English | MEDLINE | ID: mdl-36692477

ABSTRACT

Cardiolipins are phospholipids that are central to proper mitochondrial functioning. Because mitochondria play crucial roles in differentiation, development, and maturation, we would also expect cardiolipin to play major roles in these processes. Indeed, cardiolipin has been implicated in the mechanism of three human diseases that affect young infants, implying developmental abnormalities. In this review, we will: (1) Review the biology of cardiolipin; (2) Outline the evidence for essential roles of cardiolipin during organismal development, including embryogenesis and cell maturation in vertebrate organisms; (3) Place the role(s) of cardiolipin during embryogenesis within the larger context of the roles of mitochondria in development; and (4) Suggest avenues for future research.


Subject(s)
Cardiolipins , Mitochondria , Animals , Humans , Cell Differentiation
10.
EMBO J ; 41(16): e111834, 2022 08 16.
Article in English | MEDLINE | ID: mdl-35912455

ABSTRACT

How cellular cues alter the mitochondrial proteome and impact the composition of mitochondrial proteins remains poorly understood. In this issue of The EMBO Journal, Patron et al (2022) identify TMBIM5 as an important link between calcium homeostasis, proton motive force, and mitochondrial proteolysis, by which the organelle can modify its protein composition. The results may be crucial for our understanding of the plasticity of mitochondria.


Subject(s)
Mitochondria , Proteostasis , Mitochondria/metabolism , Mitochondrial Proteins/genetics , Mitochondrial Proteins/metabolism , Proteolysis , Proteome/metabolism
11.
Front Cell Dev Biol ; 10: 867175, 2022.
Article in English | MEDLINE | ID: mdl-35531097

ABSTRACT

Mammalian spermatogenesis is associated with the transient appearance of condensed mitochondria, a singularity of germ cells with unknown function. Using proteomic analysis, respirometry, and electron microscopy with tomography, we studied the development of condensed mitochondria. Condensed mitochondria arose from orthodox mitochondria during meiosis by progressive contraction of the matrix space, which was accompanied by an initial expansion and a subsequent reduction of the surface area of the inner membrane. Compared to orthodox mitochondria, condensed mitochondria respired more actively, had a higher concentration of respiratory enzymes and supercomplexes, and contained more proteins involved in protein import and expression. After the completion of meiosis, the abundance of condensed mitochondria declined, which coincided with the onset of the biogenesis of acrosomes. Immuno-electron microscopy and the analysis of sub-cellular fractions suggested that condensed mitochondria or their fragments were translocated into the lumen of the acrosome. Thus, it seems condensed mitochondria are formed from orthodox mitochondria by extensive transformations in order to support the formation of the acrosomal matrix.

12.
PLoS One ; 17(4): e0259752, 2022.
Article in English | MEDLINE | ID: mdl-35452450

ABSTRACT

Cardiolipin is known to interact with bacterial and mitochondrial proteins and protein complexes. Unlike in Escherichia coli and Saccharomyces cerevisiae, the synthesis of cardiolipin is essential for growth of Trypanosoma brucei parasites in culture. Inhibition of cardiolipin production has been shown to result in major changes in the T. brucei proteome and energy metabolism, with CLDP43, a mitochondrial protein containing a StaR-related lipid transfer (START)-like domain, being depleted in a cardiolipin-dependent way. We now show that in T. brucei procyclic forms lacking CLDP43, cardiolipin metabolism and mitochondrial function are affected. Using quantitative and qualitative lipid analyses, we found that while steady-state levels of cardiolipin were elevated in CLDP43 knock-out parasites compared to parental cells, de novo formation of cardiolipin was down-regulated. In addition, depletion of CLDP43 resulted in partial loss of mitochondrial membrane potential and decreased ATP production via substrate level phosphorylation. Recombinant CLDP43 was found to bind cardiolipin and phosphatidic acid in lipid overlay experiments, suggesting that it may be involved in transport or synthesis of cardiolipin or its precursors in T. brucei.


Subject(s)
Trypanosoma brucei brucei , Cardiolipins/metabolism , Mitochondria/metabolism , Mitochondrial Proteins/genetics , Mitochondrial Proteins/metabolism , Protozoan Proteins/metabolism , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/metabolism , Trypanosoma brucei brucei/genetics
13.
J Biol Chem ; 298(3): 101685, 2022 03.
Article in English | MEDLINE | ID: mdl-35131264

ABSTRACT

Most mammalian phospholipids contain a saturated fatty acid at the sn-1 carbon atom and an unsaturated fatty acid at the sn-2 carbon atom of the glycerol backbone group. While the sn-2 linked chains undergo extensive remodeling by deacylation and reacylation (Lands cycle), it is not known how the composition of saturated fatty acids is controlled at the sn-1 position. Here, we demonstrate that lysophosphatidylglycerol acyltransferase 1 (LPGAT1) is an sn-1 specific acyltransferase that controls the stearate/palmitate ratio of phosphatidylethanolamine (PE) and phosphatidylcholine. Bacterially expressed murine LPGAT1 transferred saturated acyl-CoAs specifically into the sn-1 position of lysophosphatidylethanolamine (LPE) rather than lysophosphatidylglycerol and preferred stearoyl-CoA over palmitoyl-CoA as the substrate. In addition, genetic ablation of LPGAT1 in mice abolished 1-LPE:stearoyl-CoA acyltransferase activity and caused a shift from stearate to palmitate species in PE, dimethyl-PE, and phosphatidylcholine. Lysophosphatidylglycerol acyltransferase 1 KO mice were leaner and had a shorter life span than their littermate controls. Finally, we show that total lipid synthesis was reduced in isolated hepatocytes of LPGAT1 knockout mice. Thus, we conclude that LPGAT1 is an sn-1 specific LPE acyltransferase that controls the stearate/palmitate homeostasis of PE and the metabolites of the PE methylation pathway and that LPGAT1 plays a central role in the regulation of lipid biosynthesis with implications for body fat content and longevity.


Subject(s)
Acyltransferases , Palmitates , Phosphatidylcholines , Stearates , Acyltransferases/metabolism , Animals , Carbon , Fatty Acids , Mammals/metabolism , Mice , Mice, Knockout , Palmitates/metabolism , Phosphatidylcholines/metabolism , Phosphatidylethanolamines , Stearates/metabolism
14.
J Inherit Metab Dis ; 45(1): 51-59, 2022 01.
Article in English | MEDLINE | ID: mdl-34611930

ABSTRACT

Barth syndrome is a multisystem disorder caused by an abnormal metabolism of the mitochondrial lipid cardiolipin. In this review, we discuss physical properties, biosynthesis, membrane assembly, and function of cardiolipin. We hypothesize that cardiolipin reduces packing stress in the inner mitochondrial membrane, which arises as a result of protein crowding. According to this hypothesis, patients with Barth syndrome are unable to meet peak energy demands because they fail to concentrate the proteins of oxidative phosphorylation to a high surface density in the inner mitochondrial membrane.


Subject(s)
Barth Syndrome/metabolism , Cardiolipins/biosynthesis , Cardiolipins/physiology , Mitochondrial Membranes/metabolism , Cardiolipins/chemistry , Humans , Mitochondria/metabolism , Oxidative Phosphorylation
15.
EMBO J ; 40(23): e108428, 2021 12 01.
Article in English | MEDLINE | ID: mdl-34661298

ABSTRACT

Mitochondrial cristae are extraordinarily crowded with proteins, which puts stress on the bilayer organization of lipids. We tested the hypothesis that the high concentration of proteins drives the tafazzin-catalyzed remodeling of fatty acids in cardiolipin, thereby reducing bilayer stress in the membrane. Specifically, we tested whether protein crowding induces cardiolipin remodeling and whether the lack of cardiolipin remodeling prevents the membrane from accumulating proteins. In vitro, the incorporation of large amounts of proteins into liposomes altered the outcome of the remodeling reaction. In yeast, the concentration of proteins involved in oxidative phosphorylation (OXPHOS) correlated with the cardiolipin composition. Genetic ablation of either remodeling or biosynthesis of cardiolipin caused a substantial drop in the surface density of OXPHOS proteins in the inner membrane of the mouse heart and Drosophila flight muscle mitochondria. Our data suggest that OXPHOS protein crowding induces cardiolipin remodelling and that remodeled cardiolipin supports the high concentration of these proteins in the inner mitochondrial membrane.


Subject(s)
Acyltransferases/physiology , Cardiolipins/metabolism , Mitochondria, Heart/metabolism , Mitochondria, Muscle/metabolism , Mitochondrial Membranes/metabolism , Oxidative Phosphorylation , Proteins/metabolism , Animals , Cardiolipins/chemistry , Cardiolipins/genetics , Drosophila melanogaster , Fatty Acids/metabolism , Female , Liposomes/metabolism , Male , Mice , Mice, Inbred C57BL , Mice, Knockout , Oxidation-Reduction , Saccharomyces cerevisiae
16.
Proc Natl Acad Sci U S A ; 118(29)2021 07 20.
Article in English | MEDLINE | ID: mdl-34272288

ABSTRACT

KdpFABC is an oligomeric K+ transport complex in prokaryotes that maintains ionic homeostasis under stress conditions. The complex comprises a channel-like subunit (KdpA) from the superfamily of K+ transporters and a pump-like subunit (KdpB) from the superfamily of P-type ATPases. Recent structural work has defined the architecture and generated contradictory hypotheses for the transport mechanism. Here, we use substrate analogs to stabilize four key intermediates in the reaction cycle and determine the corresponding structures by cryogenic electron microscopy. We find that KdpB undergoes conformational changes consistent with other representatives from the P-type superfamily, whereas KdpA, KdpC, and KdpF remain static. We observe a series of spherical densities that we assign as K+ or water and which define a pathway for K+ transport. This pathway runs through an intramembrane tunnel in KdpA and delivers ions to sites in the membrane domain of KdpB. Our structures suggest a mechanism where ATP hydrolysis is coupled to K+ transfer between alternative sites in KdpB, ultimately reaching a low-affinity site where a water-filled pathway allows release of K+ to the cytoplasm.


Subject(s)
Adenosine Triphosphatases/chemistry , Adenosine Triphosphatases/metabolism , Cation Transport Proteins/chemistry , Cation Transport Proteins/metabolism , Escherichia coli Proteins/chemistry , Escherichia coli Proteins/metabolism , Escherichia coli/enzymology , Membrane Proteins/chemistry , Membrane Proteins/metabolism , Adenosine Triphosphatases/genetics , Binding Sites , Cation Transport Proteins/genetics , Escherichia coli/chemistry , Escherichia coli/genetics , Escherichia coli/metabolism , Escherichia coli Proteins/genetics , Ion Transport , Membrane Proteins/genetics , Models, Molecular , Operon , Potassium/metabolism
17.
J Biol Chem ; 297(1): 100813, 2021 07.
Article in English | MEDLINE | ID: mdl-34023384

ABSTRACT

Niemann-Pick C (NPC) is an autosomal recessive disorder characterized by mutations in the NPC1 or NPC2 genes encoding endolysosomal lipid transport proteins, leading to cholesterol accumulation and autophagy dysfunction. We have previously shown that enrichment of NPC1-deficient cells with the anionic lipid lysobisphosphatidic acid (LBPA; also called bis(monoacylglycerol)phosphate) via treatment with its precursor phosphatidylglycerol (PG) results in a dramatic decrease in cholesterol storage. However, the mechanisms underlying this reduction are unknown. In the present study, we showed using biochemical and imaging approaches in both NPC1-deficient cellular models and an NPC1 mouse model that PG incubation/LBPA enrichment significantly improved the compromised autophagic flux associated with NPC1 disease, providing a route for NPC1-independent endolysosomal cholesterol mobilization. PG/LBPA enrichment specifically enhanced the late stages of autophagy, and effects were mediated by activation of the lysosomal enzyme acid sphingomyelinase. PG incubation also led to robust and specific increases in LBPA species with polyunsaturated acyl chains, potentially increasing the propensity for membrane fusion events, which are critical for late-stage autophagy progression. Finally, we demonstrated that PG/LBPA treatment efficiently cleared cholesterol and toxic protein aggregates in Purkinje neurons of the NPC1I1061T mouse model. Collectively, these findings provide a mechanistic basis supporting cellular LBPA as a potential new target for therapeutic intervention in NPC disease.


Subject(s)
Autophagy , Cholesterol/metabolism , Intracellular Signaling Peptides and Proteins/deficiency , Lysophospholipids/metabolism , Lysosomes/metabolism , Monoglycerides/metabolism , Animals , Autophagy/drug effects , Endosomes/metabolism , Fibroblasts/drug effects , Fibroblasts/metabolism , HeLa Cells , Homeostasis/drug effects , Humans , Intracellular Signaling Peptides and Proteins/metabolism , Lysosomes/drug effects , Mice, Inbred C57BL , Mice, Transgenic , Models, Biological , Mutation/genetics , Neural Stem Cells/drug effects , Neural Stem Cells/metabolism , Niemann-Pick C1 Protein , Niemann-Pick Disease, Type C/genetics , Phosphatidylglycerols/pharmacology , Purkinje Cells/drug effects , Purkinje Cells/metabolism , Sequestosome-1 Protein/metabolism , Sphingomyelin Phosphodiesterase/metabolism
18.
Circulation ; 143(19): 1894-1911, 2021 05 11.
Article in English | MEDLINE | ID: mdl-33793303

ABSTRACT

BACKGROUND: Mutations in tafazzin (TAZ), a gene required for biogenesis of cardiolipin, the signature phospholipid of the inner mitochondrial membrane, causes Barth syndrome (BTHS). Cardiomyopathy and risk of sudden cardiac death are prominent features of BTHS, but the mechanisms by which impaired cardiolipin biogenesis causes cardiac muscle weakness and arrhythmia are poorly understood. METHODS: We performed in vivo electrophysiology to define arrhythmia vulnerability in cardiac-specific TAZ knockout mice. Using cardiomyocytes derived from human induced pluripotent stem cells and cardiac-specific TAZ knockout mice as model systems, we investigated the effect of TAZ inactivation on Ca2+ handling. Through genome editing and pharmacology, we defined a molecular link between TAZ mutation and abnormal Ca2+ handling and contractility. RESULTS: A subset of mice with cardiac-specific TAZ inactivation developed arrhythmias, including bidirectional ventricular tachycardia, atrial tachycardia, and complete atrioventricular block. Compared with wild-type controls, BTHS-induced pluripotent stem cell-derived cardiomyocytes had increased diastolic Ca2+ and decreased Ca2+ transient amplitude. BTHS-induced pluripotent stem cell-derived cardiomyocytes had higher levels of mitochondrial and cellular reactive oxygen species than wild-type controls, which activated CaMKII (Ca2+/calmodulin-dependent protein kinase II). Activated CaMKII phosphorylated the RYR2 (ryanodine receptor 2) on serine 2814, increasing Ca2+ leak through RYR2. Inhibition of this reactive oxygen species-CaMKII-RYR2 pathway through pharmacological inhibitors or genome editing normalized aberrant Ca2+ handling in BTHS-induced pluripotent stem cell-derived cardiomyocytes and improved their contractile function. Murine Taz knockout cardiomyocytes also exhibited elevated diastolic Ca2+ and decreased Ca2+ transient amplitude. These abnormalities were ameliorated by Ca2+/calmodulin-dependent protein kinase II or reactive oxygen species inhibition. CONCLUSIONS: This study identified a molecular pathway that links TAZ mutation with abnormal Ca2+ handling and decreased cardiomyocyte contractility. This pathway may offer therapeutic opportunities to treat BTHS and potentially other diseases with elevated mitochondrial reactive oxygen species production.


Subject(s)
Barth Syndrome/genetics , Calcium-Calmodulin-Dependent Protein Kinase Type 2/metabolism , Calcium/metabolism , Reactive Oxygen Species/metabolism , Animals , Barth Syndrome/physiopathology , Humans , Mice , Mice, Knockout
19.
Biochim Biophys Acta Bioenerg ; 1862(1): 148305, 2021 01 01.
Article in English | MEDLINE | ID: mdl-32916174

ABSTRACT

The inner membrane of mitochondria is known for its low lipid-to-protein ratio. Calculations based on the size and the concentration of the principal membrane components, suggest about half of the hydrophobic volume of the membrane is occupied by proteins. Such high degree of crowding is expected to strain the hydrophobic coupling between proteins and lipids unless stabilizing mechanisms are in place. Both protein supercomplexes and cardiolipin are likely to be critical for the integrity of the inner mitochondrial membrane because they reduce the energy penalty of crowding.


Subject(s)
Mitochondria/metabolism , Mitochondrial Membranes/metabolism , Mitochondrial Proteins/metabolism , Animals , Humans
20.
FEBS Lett ; 595(3): 415-432, 2021 02.
Article in English | MEDLINE | ID: mdl-33112430

ABSTRACT

Barth syndrome (BTHS) is a rare X-linked genetic disorder caused by mutations in the gene encoding the transacylase tafazzin and characterized by loss of cardiolipin and severe cardiomyopathy. Mitochondrial oxidants have been implicated in the cardiomyopathy in BTHS. Eleven mitochondrial sites produce superoxide/hydrogen peroxide (H2 O2 ) at significant rates. Which of these sites generate oxidants at excessive rates in BTHS is unknown. Here, we measured the maximum capacity of superoxide/H2 O2 production from each site and the ex vivo rate of superoxide/H2 O2 production in the heart and skeletal muscle mitochondria of the tafazzin knockdown mice (tazkd) from 3 to 12 months of age. Despite reduced oxidative capacity, superoxide/H2 O2 production was indistinguishable between tazkd mice and wild-type littermates. These observations raise questions about the involvement of mitochondrial oxidants in BTHS pathology.


Subject(s)
Acyltransferases/genetics , Barth Syndrome/genetics , Mitochondria, Heart/enzymology , Mitochondria, Muscle/enzymology , Muscle, Skeletal/enzymology , Myocardium/enzymology , Acyltransferases/deficiency , Animals , Barth Syndrome/enzymology , Barth Syndrome/pathology , Cardiolipins/metabolism , Disease Models, Animal , Electron Transport Chain Complex Proteins , Gene Expression , Humans , Hydrogen Peroxide/metabolism , Mice , Mice, Knockout , Mitochondria, Heart/pathology , Mitochondria, Muscle/pathology , Muscle, Skeletal/pathology , Myocardium/pathology , NAD/metabolism , Oxygen Consumption/genetics , Superoxides/metabolism
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