Mitochondrial Calcium Uniporter Research Articles

Layer-specific mitochondrial diversity across hippocampal CA2 dendrites.

CA2 is an understudied subregion of the hippocampus that is critical for social memory. Previous studies identified multiple components of the mitochondrial calcium uniporter (MCU) complex as selectively enriched in CA2. The MCU complex regulates calcium entry into mitochondria, which in turn regulates mitochondrial transport and localization to active synapses. We found that MCU is strikingly enriched in CA2 distal apical dendrites, precisely where CA2 neurons receive entorhinal cortical input carrying social information. Furthermore, MCU-enriched mitochondria in CA2 distal dendrites are larger compared to mitochondria in CA2 proximal apical dendrites and neighboring CA1 apical dendrites, which was confirmed in CA2 with genetically labeled mitochondria and electron microscopy. MCU overexpression in neighboring CA1 led to a preferential localization of MCU in the proximal dendrites of CA1 compared to the distal dendrites, an effect not seen in CA2. Our findings demonstrate that mitochondria are molecularly and structurally diverse across hippocampal cell types and circuits, and suggest that MCU can be differentially localized within dendrites, possibly to meet local energy demands.

IGF-1 boosts mitochondrial function by a Ca2+ uptake-dependent mechanism in cultured human and rat cardiomyocytes.

A physiological increase in cardiac workload results in adaptive cardiac remodeling, characterized by increased oxidative metabolism and improvements in cardiac performance. Insulin-like growth factor-1 (IGF-1) has been identified as a critical regulator of physiological cardiac growth, but its precise role in cardiometabolic adaptations to physiological stress remains unresolved. Mitochondrial calcium (Ca2+) handling has been proposed to be required for sustaining key mitochondrial dehydrogenase activity and energy production during increased workload conditions, thus ensuring the adaptive cardiac response. We hypothesized that IGF-1 enhances mitochondrial energy production through a Ca2+-dependent mechanism to ensure adaptive cardiomyocyte growth. We found that stimulation with IGF-1 resulted in increased mitochondrial Ca2+ uptake in neonatal rat ventricular myocytes and human embryonic stem cell-derived cardiomyocytes, estimated by fluorescence microscopy and indirectly by a reduction in the pyruvate dehydrogenase phosphorylation. We showed that IGF-1 modulated the expression of mitochondrial Ca2+ uniporter (MCU) complex subunits and increased the mitochondrial membrane potential; consistent with higher MCU-mediated Ca2+ transport. Finally, we showed that IGF-1 improved mitochondrial respiration through a mechanism dependent on MCU-mediated Ca2+ transport. In conclusion, IGF-1-induced mitochondrial Ca2+ uptake is required to boost oxidative metabolism during cardiomyocyte adaptive growth.

MARS2 drives metabolic switch of non-small-cell lung cancer cells via interaction with MCU

Mitochondrial methionyl-tRNA synthetase (MARS2) canonically mediates the formation of fMet-tRNAifMet for mitochondrial translation initiation. Mitochondrial calcium uniporter (MCU) is a major gate of Ca2+ flux from cytosol into the mitochondrial matrix. We found that MARS2 interacts with MCU and stimulates mitochondrial Ca2+ influx. Methionine binding to MARS2 would act as a molecular switch that regulates MARS2-MCU interaction. Endogenous knockdown of MARS2 attenuates mitochondrial Ca2+ influx and induces p53 upregulation through the Ca2+-dependent CaMKII/CREB signaling. Subsequently, metabolic rewiring from glycolysis into pentose phosphate pathway is triggered and cellular reactive oxygen species level decreases. This metabolic switch induces inhibition of epithelial-mesenchymal transition (EMT) via cellular redox regulation. Expression of MARS2 is regulated by ZEB1 transcription factor in response to Wnt signaling. Our results suggest the mechanisms of mitochondrial Ca2+ uptake and metabolic control of cancer that are exerted by the key factors of the mitochondrial translational machinery and Ca2+ homeostasis.

Acquired disorders of mitochondrial metabolism and dynamics in pulmonary arterial hypertension.

Pulmonary arterial hypertension (PAH) is an orphan disease of the cardiopulmonary unit that reflects an obstructive pulmonary vasculopathy and presents with hypertrophy, inflammation, fibrosis, and ultimately failure of the right ventricle (RVF). Despite treatment using pulmonary hypertension (PH)-targeted therapies, persistent functional impairment reduces the quality of life for people with PAH and death from RVF occurs in approximately 40% of patients within 5 years of diagnosis. PH-targeted therapeutics are primarily vasodilators and none, alone or in combination, are curative. This highlights a need to therapeutically explore molecular targets in other pathways that are involved in the pathogenesis of PAH. Several candidate pathways in PAH involve acquired mitochondrial dysfunction. These mitochondrial disorders include: 1) a shift in metabolism related to increased expression of pyruvate dehydrogenase kinase and pyruvate kinase, which together increase uncoupled glycolysis (Warburg metabolism); 2) disruption of oxygen-sensing related to increased expression of hypoxia-inducible factor 1α, resulting in a state of pseudohypoxia; 3) altered mitochondrial calcium homeostasis related to impaired function of the mitochondrial calcium uniporter complex, which elevates cytosolic calcium and reduces intramitochondrial calcium; and 4) abnormal mitochondrial dynamics related to increased expression of dynamin-related protein 1 and its binding partners, such as mitochondrial dynamics proteins of 49kDa and 51kDa, and depressed expression of mitofusin 2, resulting in increased mitotic fission. These acquired mitochondrial abnormalities increase proliferation and impair apoptosis in most pulmonary vascular cells (including endothelial cells, smooth muscle cells and fibroblasts). In the RV, Warburg metabolism and induction of glutaminolysis impairs bioenergetics and promotes hypokinesis, hypertrophy, and fibrosis. This review will explore our current knowledge of the causes and consequences of disordered mitochondrial function in PAH.

Modulation of the mitochondrial Ca2+ uniporter complex subunit expression by different shear stress patterns in vascular endothelial cells

Mitochondrial calcium (m Ca2+ ) uptake occurs via the Mitochondrial Ca2+ Uniporter (MCU) complex and plays a critical role in mitochondrial dynamics, mitophagy, and apoptosis. MCU complex activity is in part modulated by the expression of its regulatory subunits. Cardiovascular disease models demonstrated altered gene/protein expression of one or multiple subunits in different cells, including vascular endothelial cells (ECs). MCU complex activity was found necessary for stable flow (s-flow)-induced mitophagy and promotion of an atheroprotective EC phenotype. Disturbed flow (d-flow) is known to lead to an atheroprone phenotype. Despite the role of MCU in flow-regulated EC function, flow-induced alterations in MCU complex subunit expression are currently unknown. We exposed cultured human ECs to atheroprotective (steady shear stress, SS) or atheroprone flow (oscillatory shear stress, OS) and measured mRNA and protein levels of the MCU complex members. SS and OS differentially modulated subunit expression at gene/protein levels. Protein expression changes of the core MCU, m Ca2+ uptake 1 (MICU1) and MCU regulator 1 (MCUR1) subunits in SS- and OS-exposed, compared to static, ECs suggested an enhanced m Ca2+ influx under each flow and a potential contribution to EC dysfunction under OS. In silico analysis of a single-cell RNA-sequencing dataset was employed to extract transcript values of MCU subunits in mouse carotid ECs from regions exposed to s-flow or d-flow. Mcu and Mcur1 genes showed significant differences in expression after prolonged exposure to each flow. The differential expression of MCU complex subunits indicated a tight regulation of the complex activity under physiological and pathological hemodynamic conditions.

Physiological significance of tissue-specific MICU regulation of mitochondrial calcium uptake.

The mitochondrial calcium uniporter is a multi-subunit calcium channel that imports Ca2+ into mitochondria. Its MICU subunits (MICU1, MICU2, and the neuron-specific MICU3) gate the channel by blocking the pore in low Ca2+. Upon local Ca2+ elevation, Ca2+ binds to MICUs to cause MICU unblock, thus opening the pore so Ca2+ can permeate. Previous work using cell lines suggests that the uniporter in mammalian cells is exclusively regulated by a MICU1-MICU2 heterodimer. However, we show here that multiple types of electrically excitable cells, including skeletal muscle and cardiac tissues, can also possess a MICU1-MICU1 homodimer or virtually no MICUs. Kinetic analyses demonstrate that MICU1 has a higher Ca2+ affinity than MICU2, and that without MICUs the uniporter is constitutively open. As a result, uniporters with the MICU1-1 homodimer or no MICUs exhibit higher transport activities, leading to mitochondria accumulating much higher levels of matrix Ca2+. Using a Seahorse assay, we show that cells with MICU1-1 or no MICUs have impaired basal oxidative phosphorylation, likely due to increased ROS and damaged respiratory-complex proteins, including NDUFS3 and COX2. These cells, moreover, are highly susceptible to apoptosis. The disadvantage of employing MICU1-1 or omitting MICUs, however, accompanies strong physiological benefits. We show that in response to intracellular Ca2+ signals, these mitochondria import more Ca2+ and consequently produce more ATP, thus better supplying the energy required for the cellular processes initiated by the Ca2+ signals. In conclusion, this work reveals that tissues can manipulate their mitochondrial calcium uptake properties to suit their unique physiological needs by customizing their MICU regulation of the mitochondrial calcium uniporter.

Molecular and cellular mechanisms underlying tissue-specific MICU-dimer assembly in the mitochondrial calcium uniporter complex.

The mitochondria calcium uniporter is a multi-subunit Ca2+ channel that mediates mitochondrial Ca2+ uptake. It regulates multiple physiological processes including oxidative phosphorylation, intracellular Ca2+ signaling, and apoptosis. The uniporter is activated by Ca2+—It remains closed at resting cellular [Ca2+] and becomes open when stimulated by intracellular Ca2+ signals. Previous work suggests that in non-neuronal cells, such Ca2+ activation is exclusively mediated by a MICU1-2 heterodimer. Here, we find that kidney and skeletal muscle uniporters also complex with a MICU1-1 homodimer. Kinetic analyses show that the uniporter is more readily activated by Ca2+ when complexed with MICU1-1 (K0.5=0.6 µM) than MICU1-2 (K0.5=1.2 µM). Thus, tissues can create specific mitochondrial Ca2+ uptake properties by varying the population of MICU1-1 and MICU1-2. Here, we demonstrate two findings to establish the mechanisms by which cells control the relative abundance of these MICU dimers. Firstly, MICU1 has a >50-fold tendency to dimerize with MICU1 than MICU2; Secondly, mitochondria import MICU2 faster than MICU1 to maintain an excess of MICU2 monomers while the MICU1 monomer is depleted. In several types of cells, due to this MICU2 excess in mitochondria, an imported MICU1 monomer can only encounter MICU2 to form MICU1-2 heterodimers. Other types of cells can produce MICU1-1 by importing MICU1 slightly faster so MICU1 could form homodimers due to its higher tendency to homodimerize. Contradicting previous studies, we show that a conserved MICU intersubunit disulfide is not required for the MICUs to regulate uniporter transport. Instead, this disulfide functions to protect properly assembled MICU dimers from proteolytic degradation by YME1L1. These results shed light on the molecular and cellular mechanisms by which different types of cells can produce unique mitochondrial Ca2+ uptake properties to fulfill their unique physiological functions.

The effect of regulating MCU expression on experimental ischemic brain injury

Mitochondrial calcium uniporter (MCU) is a critical channel for Ca2+ influx into mitochondria. The present study aimed to determine if MCU knockdown has beneficial effects on ischemic brain injury and to explore the underlying mechanisms. The present study demonstrated that MCU knockdown but not total knockout (KO) attenuated ischemia infarction volume and primary cortical neuronal cells' ischemic damage. MCU knockdown maintained mitochondrial ultrastructure, alleviated calcium overload, and reduced mitochondrial apoptosis. Moreover, MCU knockdown regulated the changes of MICU1 and MICU2 after cerebral infarction, while no changes were observed in other mitochondrial calcium handling proteins. Based on metabolomics, MCU knockdown reversed middle cerebral artery occlusion (MCAO)-induced up-regulated phosphoenolpyruvate and down-regulated GDP to protect energy metabolism after cerebral infarction. Furthermore, a total of 87 and 245 differentially expressed genes (DEGs) were detected by transcriptome sequencing among WT mice, MCU KO mice and MCU knockdown mice in the MCAO model, respectively. Then, NR4A1 was identified as one of the DEGs in different MCU expressions in vivo ischemia stroke model via transcriptomic screening and genetic validation. Furthermore, MCU knockdown downregulated the ischemia-induced upregulation of NR4A1 expression. Together, this is the further evidence that the MCU knockdown exerts a protective role after cerebral infarction by promoting calcium homeostasis, inhibiting mitochondrial apoptosis and protecting energy metabolism.

Mitochondrial calcium cycling in neuronal function and neurodegeneration.

Mitochondria are essential for proper cellular function through their critical roles in ATP synthesis, reactive oxygen species production, calcium (Ca2+) buffering, and apoptotic signaling. In neurons, Ca2+ buffering is particularly important as it helps to shape Ca2+ signals and to regulate numerous Ca2+-dependent functions including neuronal excitability, synaptic transmission, gene expression, and neuronal toxicity. Over the past decade, identification of the mitochondrial Ca2+ uniporter (MCU) and other molecular components of mitochondrial Ca2+ transport has provided insight into the roles that mitochondrial Ca2+ regulation plays in neuronal function in health and disease. In this review, we discuss the many roles of mitochondrial Ca2+ uptake and release mechanisms in normal neuronal function and highlight new insights into the Ca2+-dependent mechanisms that drive mitochondrial dysfunction in neurologic diseases including epilepsy, Alzheimer's disease, Parkinson's disease, and amyotrophic lateral sclerosis. We also consider how targeting Ca2+ uptake and release mechanisms could facilitate the development of novel therapeutic strategies for neurological diseases.

Cx43 acts as a mitochondrial calcium regulator that promotes obesity by inducing the polarization of macrophages in adipose tissue

Metabolic reprogramming of macrophages initiates the polarization of pro-inflammatory macrophages that exacerbates adipocyte dysfunction and obesity. The imbalance of mitochondrial Ca2+ homeostasis impairs mitochondrial function and promotes inflammation. Connexin 43 (Cx43), a ubiquitous gap junction protein, has been demonstrated to regulate intracellular Ca2+ homeostasis. Here we explored whether macrophage Cx43 affects the obesity process by regulating the polarization of macrophage. HFD treatment induced obesity and exacerbated macrophages infiltration with upregulation of macrophages Cx43. Macrophage-specific knockout of Cx43 reduced HFD-induced obesity by alleviating inflammation in adipose tissue, with less pro-inflammatory M1 macrophage infiltration. Consistently, inhibition or knockdown of Cx43 improved palmitic acid (PA) induced mitochondrial dysfunction, as indicated by improved oxidative phosphorylation (OXPHOS), reduced formation of mitochondria-associated membranes (MAM) and mitochondrial Ca2+ overload. Mechanistically, Cx43 interacted with the mitochondrial Ca2+ uniporter (MCU) and knockdown of Cx43 alleviated PA-induced succinate dehydrogenase (SDH) oxidation by lowering MCU-mediated mitochondrial Ca2+ uptake, which then, promoting the polarization of pro-inflammatory M1 macrophages. Thus, this study identified Cx43 as a mitochondrial Ca2+ regulator that aggravates obesity via promoting macrophages polarized to M1 pro-inflammatory phenotype and suggests that Cx43 might be a promising therapeutic target antagonizing obesity.

Investigation of Cobalt(III) Cage Complexes as Inhibitors of the Mitochondrial Calcium Uniporter.

The mitochondrial calcium uniporter (MCU) mediates uptake of calcium ions (Ca2+) into the mitochondria, a process that is vital for maintaining normal cellular function. Inhibitors of the MCU, the most promising of which are dinuclear ruthenium coordination compounds, have found use as both therapeutic agents and tools for studying the importance of this ion channel. In this study, six Co3+ cage compounds with sarcophagine-like ligands were assessed for their abilities to inhibit MCU-mediated mitochondrial Ca2+ uptake. These complexes were synthesized and characterized according to literature procedures and then investigated in cellular systems for their MCU-inhibitory activities. Among these six compounds, [Co(sen)]3+ (3, sen = 5-(4-amino-2-azabutyl)-5-methyl-3,7-diaza-1,9-nonanediamine) was identified to be a potent MCU inhibitor, with IC50 values of inhibition of 160 and 180 nM in permeabilized HeLa and HEK293T cells, respectively. Furthermore, the cellular uptake of compound 3 was determined, revealing moderate accumulation in cells. Most notably, 3 was demonstrated to operate in intact cells as an MCU inhibitor. Collectively, this work presents the viability of using cobalt coordination complexes as MCU inhibitors, providing a new direction for researchers to investigate in future studies.

Mitofusin 1 and 2 differentially regulate mitochondrial function underlying Ca2+ signaling and proliferation in rat aortic smooth muscle cells

Mitochondria play a substantial role in cytosolic Ca2+ buffering and energy metabolism. We recently demonstrated that mitofusin 2 (Mfn2) regulated Ca2+ signaling by tethering mitochondria and sarcoplasmic reticulum (SR), and thus, facilitated mitochondrial function and the proliferation of vascular smooth muscle cells (VSMCs). However, the physiological role of mitofusin 1 (Mfn1) on Ca2+ signaling and mitochondrial function remains unclear. Herein, the roles of Mfn1 and Mfn2 in mitochondrial function underlying Ca2+ signaling, ATP production, and cell proliferation were examined in rat aortic smooth muscle A10 cells. Following an arginine vasopressin-induced increase in cytosolic Ca2+ concentration ([Ca2+]cyt), Mfn2 siRNA (siMfn2) reduced cytosolic Ca2+ removal and mitochondrial Ca2+ uptake. However, Mfn1 siRNA (siMfn1) attenuated mitochondrial Ca2+ uptake, facilitated Ca2+ removal from mitochondria, and resulted in increased [Ca2+]cyt, which was mediated by the downregulation of mitochondrial Ca2+ uniporter (MCU) expression and the upregulation of mitochondrial Na+/Ca2+ exchanger (NCLX) expression. Furthermore, siMfn1 increased the mitochondrial membrane potential, ATP production by adenine nucleotide translocase (ANT), and cell proliferation, whereas siMfn2 exhibited the opposite responses. In conclusion, Mfn1 modulates the expressions of MCU, NCLX, and ANT, and Mfn2 tethers mitochondria to SR, which demonstrates their different mitochondrial functions for Ca2+ signaling, ATP production, and the proliferation of VSMCs.

MCU Upregulation Overactivates Mitophagy by Promoting VDAC1 Dimerization and Ubiquitination in the Hepatotoxicity of Cadmium.

Cadmium (Cd) is a high-risk pathogenic toxin for hepatic diseases. Excessive mitophagy is a hallmark in Cd-induced hepatotoxicity. However, the underlying mechanism remains obscure. Mitochondrial calcium uniporter (MCU) is a key regulator for mitochondrial and cellular homeostasis. Here, Cd exposure upregulated MCU expression and increased mitochondrial Ca2+ uptake are found. MCU inhibition through siRNA or by Ru360 significantly attenuates Cd-induced excessive mitophagy, thereby rescues mitochondrial dysfunction and increases hepatocyte viability. Heterozygous MCU knockout mice exhibit improved liver function, ameliorated pathological damage, less mitochondrial fragmentation, and mitophagy after Cd exposure. Mechanistically, Cd upregulates MCU expression through phosphorylation activation of cAMP-response element binding protein at Ser133(CREBS133 ) and subsequent binding of MCU promoter at the TGAGGTCT, ACGTCA, and CTCCGTGATGTA regions, leading to increased MCU gene transcription. The upregulated MCU intensively interacts with voltage-dependent anion-selective channel protein 1 (VDAC1), enhances its dimerization and ubiquitination, resulting in excessive mitophagy. This study reveals a novel mechanism, through which Cd upregulates MCU to enhance mitophagy and hepatotoxicity.

The role of calcium, Akt and ERK signaling in cadmium-induced hair cell death

Exposure to heavy metals has been shown to cause damage to a variety of different tissues and cell types including hair cells, the sensory cells of our inner ears responsible for hearing and balance. Elevated levels of one such metal, cadmium, have been associated with hearing loss and shown to cause hair cell death in multiple experimental models. While the mechanisms of cadmium-induced cell death have been extensively studied in other cell types they remain relatively unknown in hair cells. We have found that calcium signaling, which is known to play a role in cadmium-induced cell death in other cell types through calmodulin and CaMKII activation as well as IP3 receptor and mitochondrial calcium uniporter mediated calcium flow, does not appear to play a significant role in cadmium-induced hair cell death. While calmodulin inhibition can partially protect hair cells this may be due to impacts on mechanotransduction activity. Removal of extracellular calcium, and inhibiting CaMKII, the IP3 receptor and the mitochondrial calcium uniporter all failed to protect against cadmium-induced hair cell death. We also found cadmium treatment increased pAkt levels in hair cells and pERK levels in supporting cells. This activation may be protective as inhibiting these pathways enhances cadmium-induced hair cell death rather than protecting cells. Thus cadmium-induced hair cell death appears distinct from cadmium-induced cell death in other cell types where calcium, Akt and ERK signaling all promote cell death.

A Fluorogenic Inhibitor of the Mitochondrial Calcium Uniporter

AbstractInhibitors of the mitochondrial calcium uniporter (MCU) are valuable tools for studying the role of mitochondrial Ca2+ in various pathophysiological conditions. In this study, a new fluorogenic MCU inhibitor, RuOCou, is presented. This compound is an analogue of the known MCU inhibitor Ru265 that contains fluorescent axial coumarin carboxylate ligands. Upon aquation of RuOCou and release of the axial coumarin ligands, a simultaneous increase in its MCU‐inhibitory activity and fluorescence intensity is observed. The fluorescence response of this compound enabled its aquation to be monitored in both HeLa cell lysates and live HeLa cells. This fluorogenic prodrug represents a potential theranostic MCU inhibitor that can be leveraged for the treatment of human diseases related to MCU activity.

A Fluorogenic Inhibitor of the Mitochondrial Calcium Uniporter.

Inhibitors of the mitochondrial calcium uniporter (MCU) are valuable tools for studying the role of mitochondrial Ca2+ in various pathophysiological conditions. In this study, a new fluorogenic MCU inhibitor, RuOCou, is presented. This compound is an analogue of the known MCU inhibitor Ru265 that contains fluorescent axial coumarin carboxylate ligands. Upon aquation of RuOCou and release of the axial coumarin ligands, a simultaneous increase in its MCU-inhibitory activity and fluorescence intensity is observed. The fluorescence response of this compound enabled its aquation to be monitored in both HeLa cell lysates and live HeLa cells. This fluorogenic prodrug represents a potential theranostic MCU inhibitor that can be leveraged for the treatment of human diseases related to MCU activity.

Mitochondrial sodium/calcium exchanger NCLX regulates glycolysis in astrocytes, impacting on cognitive performance.

Intracellular Ca2+ concentrations are strictly controlled by plasma membrane transporters, the endoplasmic reticulum, and mitochondria, in which Ca2+ uptake is mediated by the mitochondrial calcium uniporter complex (MCUc), while efflux occurs mainly through the mitochondrial Na+ /Ca2+ exchanger (NCLX). RNAseq database repository searches led us to identify the Nclx transcript as highly enriched in astrocytes when compared with neurons. To assess the role of NCLX in mouse primary culture astrocytes, we inhibited its function both pharmacologically or genetically. This resulted in re-shaping of cytosolic Ca2+ signaling and a metabolic shift that increased glycolytic flux and lactate secretion in a Ca2+ -dependent manner. Interestingly, in vivo genetic deletion of NCLX in hippocampal astrocytes improved cognitive performance in behavioral tasks, whereas hippocampal neuron-specific deletion of NCLX impaired cognitive performance. These results unveil a role for NCLX as a novel modulator of astrocytic glucose metabolism, impacting on cognition.

Mitochondrial calcium uniporter activates TFEB-driven autophagy to promote migration of breast cancer cells.

Tumor metastasis is the leading cause of death in breast cancer (BC) patients and is a complicated process. Mitochondrial calcium uniporter (MCU), a selective channel responsible for mitochondrial Ca2+ uptake, has been reported to be associated with tumorigenesis and metastasis. The molecular mechanisms of MCU contributing to the migration of BC cells are partially understood. This study investigated the role of MCU in BC cell metastasis and explored the underlying mechanism of MCU-mediated autophagy in BC cell migration. The Kaplan-Meier plotter database was used to analyze the prognostic value of MCU mRNA expression. Western blotting was used to examine the expression level of MCU in 4 paired BC and adjacent normal tissues. The cellular migration capability of BC was measured by transwell migration assay and wound healing assay. Western blotting and reverse transcription-quantitative polymerase chain reaction were performed to detect the expression levels of autophagy-related markers. The effects of MCU activation or inhibition on TFEB nuclear translocation in BC cells were detected by laser scanning confocal microscopy. Expression of MCU was found to be negatively correlated with BC patient prognosis in the Kaplan-Meier plotter database. Compared with the adjacent normal tissues, MCU was markedly up-regulated in the BC tissues. MCU overexpression promoted cellular migration, activated autophagy, and increased TFEB nuclear translocation in BC cells, whereas its knockdown produced the opposite effects. MCU activates TFEB-driven autophagy to promote BC cell metastasis and provides a potential novel therapeutic target for BC clinical intervention.

Dinuclear nitrido-bridged osmium complexes inhibit the mitochondrial calcium uniporter and protect cortical neurons against lethal oxygen-glucose deprivation.

Dysregulation of mitochondrial calcium uptake mediated by the mitochondrial calcium uniporter (MCU) is implicated in several pathophysiological conditions. Dinuclear ruthenium complexes are effective inhibitors of the MCU and have been leveraged as both tools to study mitochondrial calcium dynamics and potential therapeutic agents. In this study, we report the synthesis and characterization of Os245 ([Os2(μ-N)(NH3)8Cl2]3+) which is the osmium-containing analogue of our previously reported ruthenium-based inhibitor Ru265. This complex and its aqua-capped analogue Os245' ([Os2(μ-N)(NH3)8(OH2)2]5+) are both effective inhibitors of the MCU in permeabilized and intact cells. In comparison to the ruthenium-based inhibitor Ru265 (k obs = 4.92 × 10-3 s-1), the axial ligand exchange kinetics of Os245 are two orders of magnitude slower (k obs = 1.63 × 10-5 s-1) at 37 °C. The MCU-inhibitory properties of Os245 and Os245' are different (Os245 IC50 for MCU inhibition = 103 nM; Os245' IC50 for MCU inhibition = 2.3 nM), indicating that the axial ligands play an important role in their interactions with this channel. We further show that inhibition of the MCU by these complexes protects primary cortical neurons against lethal oxygen-glucose deprivation. When administered in vivo to mice (10 mg kg-1), Os245 and Os245' induce seizure-like behaviors in a manner similar to the ruthenium-based inhibitors. However, the onset of these seizures is delayed, a possible consequence of the slower ligand substitution kinetics for these osmium complexes. These findings support previous studies that demonstrate inhibition of the MCU is a promising therapeutic strategy for the treatment of ischemic stroke, but also highlight the need for improved drug delivery strategies to mitigate the pro-convulsant effects of this class of complexes before they can be implemented as therapeutic agents. Furthermore, the slower ligand substitution kinetics of the osmium analogues may afford new strategies for the development and modification of this class of MCU inhibitors.

Identification and Validation of a Mitochondria Calcium Uptake-Related Gene Signature for Predicting Prognosis in COAD.

Background: Mitochondrial calcium uniporter (MCU) complex has been reported to be associated with the tumor occurrence and development in varieties of malignancies. However, the role of MCU complex in colon adenocarcinoma (COAD) remains unclear. Therefore, we constructed a risk score signature based on the MCU complex members to predict the prognosis and response to immunotherapy for patients with COAD. Methods: The MCU complex-associated risk signature (MCUrisk) was constructed based on the expressions of MCU, MCUb, MCUR1, SMDT1, MICU1, MICU2, and MICU3 in COAD. The immune score, stromal score, tumor purity and estimate score were calculated by the ESTIMATE algorithm. We systematically evaluated the relationship among the MCUrisk, mutation signature, immune cell infiltration, and immune checkpoint molecules. The response to immunotherapy was quantified by the Tumor Immune Dysfunction and Exclusion (TIDE). Results: Our results showed that high score of MCUrisk was a worse factor for overall survival (OS) in COAD, and MCUrisk score was significantly higher in advanced COAD. The mutation landscape was different between the MCUrisk-high and MCUrisk-low groups, and the mutation rate of TP53 was remarkably higher in MCUrisk-high group, which strongly suggested TP53 mutation might be associated with mitochondrial calcium dyshomeostasis in COAD. Furthermore, MCUrisk score was negatively correlated with tumor mutation burden (TMB), and combining risk score and TMB as a novel index was better than TMB alone in predicting the prognosis for COAD patients. The compositions of Tregs and M0/M2 macrophages were significantly increased in MCUrisk-high group, whereas CD4+ T cells was significantly decreased in MCUrisk-high group. Consistently, the immune score was lower in MCUrisk-high group. The expression levels of immune checkpoint molecules were negatively correlated with the MCUrisk score, including CD58 and CD226. Furthermore, a lower MCUrisk score indicated better response to immunotherapy, and combining risk score and immune score was a novel indicator to precisely predict the response to immuotherapy for COAD patients. Conclusion: Altogether, a novel MCUrisk signature was constructed based on the mitochondrial calcium uptake-associated genes, and a lower MCUrisk score may predict better OS outcome and better response to immunotherapy in COAD.