Showing papers on "Mitochondrial carrier published in 2021"

Glutathione Metabolism and the Novel Role of Mitochondrial GSH in Retinal Degeneration

Abstract: Glutathione (GSH) is present ubiquitously, and its role as a crucial cellular antioxidant in tissues, including the retina, is well established. GSH’s antioxidant function arises from its ability to scavenge reactive oxygen species or to serve as an essential cofactor for GSH S-transferases and peroxidases. This review summarizes the general functions, retinal distribution, disorders linked to GSH deficiency, and the emerging role for mitochondrial GSH (mGSH) in retinal function. Though synthesized only in the cytosol, the presence of GSH in multiple cell organelles suggests the requirement for its active transport across organellar membranes. The localization and distribution of 2-oxoglutarate carrier (OGC) and dicarboxylate carrier (DIC), two recently characterized mitochondrial carrier proteins in RPE and retina, show that these transporters are highly expressed in human retinal pigment epithelium (RPE) cells and retinal layers, and their expression increases with RPE polarity in cultured cells. Depletion of mGSH levels via inhibition of the two transporters resulted in reduced mitochondrial bioenergetic parameters (basal respiration, ATP production, maximal respiration, and spare respiratory capacity) and increased RPE cell death. These results begin to reveal a critical role for mGSH in maintaining RPE bioenergetics and cell health. Thus, augmentation of mGSH pool under GSH-deficient conditions may be a valuable tool in treating retinal disorders, such as age-related macular degeneration and optic neuropathies, whose pathologies have been associated with mitochondrial dysfunction.

The versatility of plant organic acid metabolism in leaves is underpinned by mitochondrial malate-citrate exchange.

Abstract: Malate and citrate underpin the characteristic flexibility of central plant metabolism by linking mitochondrial respiratory metabolism with cytosolic biosynthetic pathways. However, the identity of mitochondrial carrier proteins that influence both processes has remained elusive. Here we show by a systems approach that DICARBOXYLATE CARRIER 2 (DIC2) facilitates mitochondrial malate-citrate exchange in vivo in Arabidopsis thaliana. DIC2 knockout (dic2-1) retards growth of vegetative tissues. In vitro and in organello analyses demonstrate that DIC2 preferentially imports malate against citrate export, which is consistent with altered malate and citrate utilisation in response to prolonged darkness of dic2-1 plants or a sudden shift to darkness of dic2-1 leaves. Furthermore, isotopic glucose tracing reveals a reduced flux towards citrate in dic2-1, which results in a metabolic diversion towards amino acid synthesis. These observations reveal the physiological function of DIC2 in mediating the flow of malate and citrate between the mitochondrial matrix and other cell compartments.

ER targeting of non-imported mitochondrial carrier proteins is dependent on the GET pathway.

Abstract: Deficiencies in mitochondrial import cause the toxic accumulation of non-imported mitochondrial precursor proteins. Numerous fates for non-imported mitochondrial precursors have been identified in budding yeast, including proteasomal destruction, deposition into protein aggregates, and mistargeting to other organelles. Amongst organelles, the ER has emerged as a key destination for a subset of non-imported mitochondrial proteins. However, how ER targeting of various types of mitochondrial proteins is achieved remains incompletely understood. Here, we show that the ER delivery of endogenous mitochondrial transmembrane proteins, especially those belonging to the SLC25A mitochondrial carrier family, is dependent on the guided entry of tail-anchored proteins (GET) complex. Without a functional GET pathway, non-imported mitochondrial proteins destined for the ER are alternatively sequestered into Hsp42-dependent protein foci. Loss of the GET pathway is detrimental to yeast cells experiencing mitochondrial import failure and prevents re-import of mitochondrial proteins from the ER via the ER-SURF pathway. Overall, this study outlines an important role for the GET complex in ER targeting of non-imported mitochondrial carrier proteins.

Structural Mechanism of Transport of Mitochondrial Carriers.

Abstract: Members of the mitochondrial carrier family [solute carrier family 25 (SLC25)] transport nucleotides, amino acids, carboxylic acids, fatty acids, inorganic ions, and vitamins across the mitochondri

The modified mitochondrial outer membrane carrier MTCH2 links mitochondrial fusion to lipogenesis.

Abstract: Mitochondrial function is integrated with cellular status through the regulation of opposing mitochondrial fusion and division events. Here we uncover a link between mitochondrial dynamics and lipid metabolism by examining the cellular role of mitochondrial carrier homologue 2 (MTCH2). MTCH2 is a modified outer mitochondrial membrane carrier protein implicated in intrinsic cell death and in the in vivo regulation of fatty acid metabolism. Our data indicate that MTCH2 is a selective effector of starvation-induced mitochondrial hyperfusion, a cytoprotective response to nutrient deprivation. We find that MTCH2 stimulates mitochondrial fusion in a manner dependent on the bioactive lipogenesis intermediate lysophosphatidic acid. We propose that MTCH2 monitors flux through the lipogenesis pathway and transmits this information to the mitochondrial fusion machinery to promote mitochondrial elongation, enhanced energy production, and cellular survival under homeostatic and starvation conditions. These findings will help resolve the roles of MTCH2 and mitochondria in tissue-specific lipid metabolism in animals.

The Mitochondrial Carnitine Acyl-carnitine Carrier (SLC25A20): Molecular Mechanisms of Transport, Role in Redox Sensing and Interaction with Drugs.

Abstract: The SLC25A20 transporter, also known as carnitine acyl-carnitine carrier (CAC), catalyzes the transport of short, medium and long carbon chain acyl-carnitines across the mitochondrial inner membrane in exchange for carnitine. The 30-year story of the protein responsible for this function started with its purification from rat liver mitochondria. Even though its 3D structure is not yet available, CAC is one of the most deeply characterized transport proteins of the inner mitochondrial membrane. Other than functional, kinetic and mechanistic data, post-translational modifications regulating the transport activity of CAC have been revealed. CAC interactions with drugs or xenobiotics relevant to human health and toxicology and the response of the carrier function to dietary compounds have been discovered. Exploiting combined approaches of site-directed mutagenesis with chemical targeting and bioinformatics, a large set of data on structure/function relationships have been obtained, giving novel information on the molecular mechanism of the transport catalyzed by this protein.

Mitochondrial copper and phosphate transporter specificity was defined early in the evolution of eukaryotes.

Abstract: The mitochondrial carrier family protein SLC25A3 transports both copper and phosphate in mammals, yet in Saccharomyces cerevisiae the transport of these substrates is partitioned across two paralogs: PIC2 and MIR1. To understand the ancestral state of copper and phosphate transport in mitochondria, we explored the evolutionary relationships of PIC2 and MIR1 orthologs across the eukaryotic tree of life. Phylogenetic analyses revealed that PIC2-like and MIR1-like orthologs are present in all major eukaryotic supergroups, indicating an ancient gene duplication created these paralogs. To link this phylogenetic signal to protein function, we used structural modeling and site-directed mutagenesis to identify residues involved in copper and phosphate transport. Based on these analyses, we generated an L175A variant of mouse SLC25A3 that retains the ability to transport copper but not phosphate. This work highlights the utility of using an evolutionary framework to uncover amino acids involved in substrate recognition by mitochondrial carrier family proteins.

Welcome to the family: Identification of the nad+ transporter of animal mitochondria as member of the solute carrier family slc25

Abstract: Subcellular compartmentation is a fundamental property of eukaryotic cells. Communication and metabolic and regulatory interconnectivity between organelles require that solutes can be transported across their surrounding membranes. Indeed, in mammals, there are hundreds of genes encoding solute carriers (SLCs) which mediate the selective transport of molecules such as nucleotides, amino acids, and sugars across biological membranes. Research over many years has identified the localization and preferred substrates of a large variety of SLCs. Of particular interest has been the SLC25 family, which includes carriers embedded in the inner membrane of mitochondria to secure the supply of these organelles with major metabolic intermediates and coenzymes. The substrate specificity of many of these carriers has been established in the past. However, the route by which animal mitochondria are supplied with NAD+ had long remained obscure. Only just recently, the existence of a human mitochondrial NAD+ carrier was firmly established. With the realization that SLC25A51 (or MCART1) represents the major mitochondrial NAD+ carrier in mammals, a long-standing mystery in NAD+ biology has been resolved. Here, we summarize the functional importance and structural features of this carrier as well as the key observations leading to its discovery.

Antioxidant Synergy of Mitochondrial Phospholipase PNPLA8/iPLA2γ with Fatty Acid–Conducting SLC25 Gene Family Transporters

Abstract: Patatin-like phospholipase domain-containing protein PNPLA8, also termed Ca2+-independent phospholipase A2γ (iPLA2γ), is addressed to the mitochondrial matrix (or peroxisomes), where it may manifest its unique activity to cleave phospholipid side-chains from both sn-1 and sn-2 positions, consequently releasing either saturated or unsaturated fatty acids (FAs), including oxidized FAs. Moreover, iPLA2γ is directly stimulated by H2O2 and, hence, is activated by redox signaling or oxidative stress. This redox activation permits the antioxidant synergy with mitochondrial uncoupling proteins (UCPs) or other SLC25 mitochondrial carrier family members by FA-mediated protonophoretic activity, termed mild uncoupling, that leads to diminishing of mitochondrial superoxide formation. This mechanism allows for the maintenance of the steady-state redox status of the cell. Besides the antioxidant role, we review the relations of iPLA2γ to lipid peroxidation since iPLA2γ is alternatively activated by cardiolipin hydroperoxides and hypothetically by structural alterations of lipid bilayer due to lipid peroxidation. Other iPLA2γ roles include the remodeling of mitochondrial (or peroxisomal) membranes and the generation of specific lipid second messengers. Thus, for example, during FA β-oxidation in pancreatic β-cells, H2O2-activated iPLA2γ supplies the GPR40 metabotropic FA receptor to amplify FA-stimulated insulin secretion. Cytoprotective roles of iPLA2γ in the heart and brain are also discussed.

Mitochondrial Carriers and Substrates Transport Network: A Lesson from Saccharomyces cerevisiae

Abstract: The yeast Saccharomyces cerevisiae is one of the most widely used model organisms for investigating various aspects of basic cellular functions that are conserved in human cells. This organism, as well as human cells, can modulate its metabolism in response to specific growth conditions, different environmental changes, and nutrient depletion. This adaptation results in a metabolic reprogramming of specific metabolic pathways. Mitochondrial carriers play a fundamental role in cellular metabolism, connecting mitochondrial with cytosolic reactions. By transporting substrates across the inner membrane of mitochondria, they contribute to many processes that are central to cellular function. The genome of Saccharomyces cerevisiae encodes 35 members of the mitochondrial carrier family, most of which have been functionally characterized. The aim of this review is to describe the role of the so far identified yeast mitochondrial carriers in cell metabolism, attempting to show the functional connections between substrates transport and specific metabolic pathways, such as oxidative phosphorylation, lipid metabolism, gluconeogenesis, and amino acids synthesis. Analysis of the literature reveals that these proteins transport substrates involved in the same metabolic pathway with a high degree of flexibility and coordination. The understanding of the role of mitochondrial carriers in yeast biology and metabolism could be useful for clarifying unexplored aspects related to the mitochondrial carrier network. Such knowledge will hopefully help in obtaining more insight into the molecular basis of human diseases.

Structural determinants of ligands recognition by the human mitochondrial basic amino acids transporter SLC25A29. Insights from molecular dynamics simulations of the c-state.

Abstract: In mitochondria, metabolic processes require the trafficking of solutes and organic molecules, such as amino acids. This task is accomplished by the Mitochondrial Carrier Family members (also known as SLC25), among which the SLC25A29 is responsible for the translocation of basic amino acids. In this regard, nitric oxide levels originated by the arginine mitochondrial catabolism have been shown to strongly affect cancer cells' metabolic status. Furthermore, the metabolic disease saccharopinuria has been linked to a mitochondrial dysregulation caused by a toxic intermediate of the lysine catabolism. In both cases, a reduction of the activity of SLC25A29 has been shown to ameliorate these pathological conditions. However, no detailed structural data are available on SLC25A29. In the present work, molecular modelling, docking and dynamics simulations have been employed to analyse the structural determinants of ligands recognition by SLC25A29 in the c-state. Results confirm and reinforce earlier predictions that Asn73, Arg160 and Glu161, and Arg257 represent the ligand contact points I, II, and III, respectively, and that Arg160, Trp204 and Arg257 form a stable interaction, likely critical for ligand binding and translocation. These results are discussed in view of the experimental data available for SLC25A29 and other homologous carriers of the same family.

Chimeric Drug Design with a Noncharged Carrier for Mitochondrial Delivery.

Abstract: Recently, it was proposed that the thiophene ring is capable of promoting mitochondrial accumulation when linked to fluorescent markers. As a noncharged group, thiophene presents several advantages from a synthetic point of view, making it easier to incorporate such a side moiety into different molecules. Herein, we confirm the general applicability of the thiophene group as a mitochondrial carrier for drugs and fluorescent markers based on a new concept of nonprotonable, noncharged transporter. We implemented this concept in a medicinal chemistry application by developing an antitumor, metabolic chimeric drug based on the pyruvate dehydrogenase kinase (PDHK) inhibitor dichloroacetate (DCA). The promising features of the thiophene moiety as a noncharged carrier for targeting mitochondria may represent a starting point for the design of new metabolism-targeting drugs.

Significance of AtMTM1 and AtMTM2 for Mitochondrial MnSOD Activation in Arabidopsis.

Abstract: The manganese (Mn) tracking factor for mitochondrial Mn superoxide dismutase (MnSOD) has been annotated as yMTM1 in yeast, which belongs to the mitochondrial carrier family. We confirmed that Arabidopsis AtMTM1 and AtMTM2 are functional homologs of yMTM1 as they can revive yeast MnSOD activity in yMTM1-mutant cells. Transient expression of AtMnSOD-3xFLAG in the AtMTM1 and AtMTM2-double mutant protoplasts confirmed that AtMTM1 and AtMTM2 are required for AtMnSOD activation. Our study revealed that AtMnSOD interacts with AtMTM1 and AtMTM2 in the mitochondria. The expression levels of AtMTM1, AtMTM2, and AtMnSOD respond positively to methyl viologen (MV) and metal stress. AtMTM1 and AtMTM2 are involved in Mn and Fe homeostasis, root length, and flowering time. Transient expression of chloroplast-destined AtMnSOD revealed that an evolutionarily conserved activation mechanism, like the chloroplastic-localized MnSOD in some algae, still exists in Arabidopsis chloroplasts. This study strengthens the proposition that AtMTM1 and AtMTM2 participate in the AtMnSOD activation and ion homeostasis.

The Interaction of Hemin, a Porphyrin Derivative, with the Purified Rat Brain 2-Oxoglutarate Carrier

Abstract: The mitochondrial 2-oxoglutarate carrier (OGC), isolated and purified from rat brain mitochondria, was reconstituted into proteoliposomes to study the interaction with hemin, a porphyrin derivative, which may result from the breakdown of heme-containing proteins and plays a key role in several metabolic pathways. By kinetic approaches, on the basis of the single binding centre gated pore mechanism, we analyzed the effect of hemin on the transport rate of OGC in uptake and efflux experiments in proteoliposomes reconstituted in the presence of the substrate 2-oxoglutarate. Overall, our experimental data fit the hypothesis that hemin operates a competitive inhibition in the 0.5-10 µM concentration range. As a consequence of the OGC inhibition, the malate/aspartate shuttle might be impaired, causing an alteration of mitochondrial function. Hence, considering that the metabolism of porphyrins implies both cytoplasmic and mitochondrial processes, OGC may participate in the regulation of porphyrin derivatives availability and the related metabolic pathways that depend on them (such as oxidative phosphorylation and apoptosis). For the sake of clarity, a simplified model based on induced-fit molecular docking supported the in vitro transport assays findings that hemin was as good as 2-oxoglutarate to bind the carrier by engaging specific ionic hydrogen bond interactions with a number of key residues known for participating in the similarly located mitochondrial carrier substrate binding site.

Molecular mechanism of thiamine pyrophosphate import into mitochondria: a molecular simulation study.

Abstract: The import of thiamine pyrophosphate (TPP) through both mitochondrial membranes was studied using a total of 3-µs molecular dynamics simulations. Regarding the translocation through the mitochondrial outer membrane, our simulations support the conjecture that TPP uses the voltage-dependent anion channel, the major pore of this membrane, for its passage to the intermembrane space, as its transport presents significant analogies with that used by other metabolites previously studied, in particular with ATP. As far as passing through the mitochondrial inner membrane is concerned, our simulations show that the specific carrier of TPP has a single binding site that becomes accessible, through an alternating access mechanism. The preference of this transporter for TPP can be rationalized mainly by three residues located in the binding site that differ from those identified in the ATP/ADP carrier, the most studied member of the mitochondrial carrier family. The simulated transport mechanism of TPP highlights the essential role, at the energetic level, of the contributions coming from the formation and breakage of two networks of salt bridges, one on the side of the matrix and the other on the side of the intermembrane space, as well as the interactions, mainly of an ionic nature, formed by TPP upon its binding. The energy contribution provided by the cytosolic network establishes a lower barrier than that of the matrix network, which can be explained by the lower interaction energy of TPP on the matrix side or possibly a uniport activity.