Crosstalk between adenine nucleotide transporter and mitochondrial swelling: experimental and computational approaches.

Mitochondrial metabolism and function are modulated by changes in matrix Ca2+. Small increases in the matrix Ca2+ stimulate mitochondrial bioenergetics, whereas excessive Ca2+ leads to cell death by causing massive matrix swelling and impairing the structural and functional integrity of mitochondria. Sustained opening of the non-selective mitochondrial permeability transition pores (PTP) is the main mechanism responsible for mitochondrial Ca2+ overload that leads to mitochondrial dysfunction and cell death. Recent studies suggest the existence of two or more types of PTP, and adenine nucleotide translocator (ANT) and FOF1-ATP synthase were proposed to form the PTP independent of each other. Here, we elucidated the role of ANT in PTP opening by applying both experimental and computational approaches. We first developed and corroborated a detailed model of the ANT transport mechanism including the matrix (ANTM), cytosolic (ANTC), and pore (ANTP) states of the transporter. Then, the ANT model was incorporated into a simple, yet effective, empirical model of mitochondrial bioenergetics to ascertain the point when Ca2+ overload initiates PTP opening via an ANT switch-like mechanism activated by matrix Ca2+ and is inhibited by extra-mitochondrial ADP. We found that encoding a heterogeneous Ca2+ response of at least three types of PTPs, weakly, moderately, and strongly sensitive to Ca2+, enabled the model to simulate Ca2+ release dynamics observed after large boluses were administered to a population of energized cardiac mitochondria. Thus, this study demonstrates the potential role of ANT in PTP gating and proposes a novel mechanism governing the cryptic nature of the PTP phenomenon.

Elucidating the contribution of mitochondrial glutathione to ferroptosis in cardiomyocytes.

Ferroptosis is a programmed iron-dependent cell death associated with peroxidation of lipids particularly, phospholipids. Several studies suggested a possible contribution of mitochondria to ferroptosis although the mechanisms underlying mitochondria-mediated ferroptotic pathways remain elusive. Reduced glutathione (GSH) is a central player in ferroptosis that is required for glutathione peroxidase 4 to eliminate oxidized phospholipids. Mitochondria do not produce GSH, and although the transport of GSH to mitochondria is not fully understood, two carrier proteins, the dicarboxylate carrier (DIC, SLC25A10) and the oxoglutarate carrier (OGC, SLC25A11) have been suggested to participate in GSH transport. Here, we elucidated the role of DIC and OGC as well as mitochondrial bioenergetics in ferroptosis in H9c2 cardioblasts. Results showed that mitochondria are highly sensitive to ferroptotic stimuli displaying fragmentation, and lipid peroxidation shortly after the onset of ferroptotic stimulus. Inhibition of electron transport chain complexes and oxidative phosphorylation worsened RSL3-induced ferroptosis. LC-MS/MS analysis revealed a dramatic increase in the levels of pro-ferroptotic oxygenated phosphatidylethanolamine species in mitochondria in response to RSL3 (ferroptosis inducer) and cardiac ischemia-reperfusion. Inhibition of DIC and OGC aggravated ferroptosis and increased mitochondrial ROS, membrane depolarization, and GSH depletion. Dihydrolipoic acid, an essential cofactor for several mitochondrial multienzyme complexes, attenuated ferroptosis and induced direct reduction of pro-ferroptotic peroxidized phospholipids to hydroxy-phospholipids in vitro. In conclusion, we suggest that ferroptotic stimuli diminishes mitochondrial bioenergetics and stimulates GSH depletion and glutathione peroxidase 4 inactivation leading to ferroptosis.

Acetylation of Mitochondrial Proteins in the Heart: The Role of SIRT3.

A growing number of studies have demonstrated the role of post-translational modifications of proteins, particularly acetylation, in human diseases including neurodegenerative and cardiovascular diseases, diabetes, cancer, and in aging. Acetylation of mitochondrial proteins has been shown to be involved in the pathogenesis of cardiac diseases such as myocardial infarction (ischemia-reperfusion) and heart failure. Indeed, over 60% of mitochondrial proteins contain acetylation sites, and most of these proteins are involved in mitochondrial bioenergetics. Mitochondrial non-enzymatic acetylation is enabled by acetyl-coenzyme A abundance and serves as the primary pathway of acetylation in mitochondria. Hence, regulation of enzymatic deacetylation becomes the most important mechanism to control acetylation/deacetylation of mitochondrial proteins. Acetylation/deacetylation of mitochondrial proteins has been regarded as a key regulator of mitochondrial metabolism and function. Proteins are deacetylated by NAD+-dependent deacetylases known as sirtuins (SIRTs). Among seven sirtuin isoforms, only SIRT3, SIRT4, and SIRT5 are localized in the mitochondria. SIRT3 is the main mitochondrial sirtuin which plays a key role in maintaining metabolic and redox balance in the mitochondria under physiological and pathological conditions. SIRT3 regulates the enzymatic activity of proteins involved in fatty acid oxidation, tricarboxylic acid cycle, electron transport chain, and oxidative phosphorylation. Although many enzymes have been identified as targets for SIRT3, cardiac-specific SIRT3 effects and regulations could differ from those in non-cardiac tissues. Therefore, it is important to elucidate the contribution of SIRT3 and mitochondrial protein acetylation/deacetylation in mitochondrial metabolism and cardiac dysfunction. Here, we summarize previous studies and provide a comprehensive analysis of the role of SIRT3 in mitochondria metabolism and bioenergetics under physiological conditions and in cardiac diseases. In addition, the review discusses mitochondrial protein acetylation as a potential target for cardioprotection.