Assembly Of Mitochondrial Complex Research Articles

HINT2 protects against pressure overload-induced cardiac remodelling through mitochondrial pathways.

Histidine triad nucleotide-binding protein 2 (HINT2) is an enzyme found in mitochondria that functions as a nucleotide hydrolase and transferase. Prior studies have demonstrated that HINT2 plays a crucial role in ischemic heart disease, but its importance in cardiac remodelling remains unknown. Therefore, the current study intends to determine the role of HINT2 in cardiac remodelling. HINT2 expression levels were found to be lower in failing hearts and hypertrophy cardiomyocytes. The mice that overexpressed HINT2 exhibited reduced myocyte hypertrophy and cardiac dysfunction in response to stress. In contrast, the deficiency of HINT2 in the heart of mice resulted in a worsening hypertrophic phenotype. Further analysis indicated that upregulated genes were predominantly associated with the oxidative phosphorylation and mitochondrial complex I pathways in HINT2-overexpressed mice after aortic banding (AB) treatment. This suggests that HINT2 increases the expression of NADH dehydrogenase (ubiquinone) flavoprotein (NDUF) genes. In cellular studies, rotenone was used to disrupt mitochondrial complex I, and the protective effect of HINT2 overexpression was nullified. Lastly, we predicted that thyroid hormone receptor beta might regulate HINT2 transcriptional activity. To conclusion, the current study showcased that HINT2 alleviates pressure overload-induced cardiac remodelling by influencing the activity and assembly of mitochondrial complex I. Thus, targeting HINT2 could be a novel therapeutic strategy for reducing cardiac remodelling.

Tissue-specific differences in the assembly of mitochondrial Complex I are revealed by a novel ENU mutation in ECSIT.

Mitochondrial Complex I assembly (MCIA) is a multi-step process that necessitates the involvement of a variety of assembly factors and chaperones to ensure that the final active enzyme is correctly assembled. The role of the assembly factor evolutionarily conserved signalling intermediate in the toll (ECSIT) pathway was studied across various murine tissues to determine its role in this process and how this varied between tissues of varying energetic demands. We hypothesized that many of the known functions of ECSIT were unhindered by the introduction of an ENU-induced mutation, while its role in Complex I assembly was affected on a tissue-specific basis. Here, we describe a mutation in the MCIA factor ECSIT that reveals tissue-specific requirements for ECSIT in Complex I assembly. MCIA is a multi-step process dependent on assembly factors that organize and arrange the individual subunits, allowing for their incorporation into the complete enzyme complex. We have identified an ENU-induced mutation in ECSIT (N209I) that exhibits a profound effect on Complex I component expression and assembly in heart tissue, resulting in hypertrophic cardiomyopathy in the absence of other phenotypes. The dysfunction of Complex I appears to be cardiac specific, leading to a loss of mitochondrial output as measured by Seahorse extracellular flux and various biochemical assays in heart tissue, while mitochondria from other tissues were unaffected. These data suggest that the mechanisms underlying Complex I assembly and activity may have tissue-specific elements tailored to the specific demands of cells and tissues. Our data suggest that tissues with high-energy demands, such as the heart, may utilize assembly factors in different ways to low-energy tissues in order to improve mitochondrial output. These data have implications for the diagnosis and treatment of various disorders of mitochondrial function as well as cardiac hypertrophy with no identifiable underlying genetic cause.

Insulin-Degrading Enzyme Interacts with Mitochondrial Ribosomes and Respiratory Chain Proteins.

Insulin-degrading enzyme (IDE) is a highly conserved metalloprotease that is mainly localized in the cytosol. Although IDE can degrade insulin and some other low molecular weight substrates efficiently, its ubiquitous expression suggests additional functions supported by experimental findings, such as a role in stress responses and cellular protein homeostasis. The translation of a long full-length IDE transcript has been reported to result in targeting to mitochondria, but the role of IDE in this compartment is unknown. To obtain initial leads on the function of IDE in mitochondria, we used a proximity biotinylation approach to identify proteins interacting with wild-type and protease-dead IDE targeted to the mitochondrial matrix. We find that IDE interacts with multiple mitochondrial ribosomal proteins as well as with proteins involved in the synthesis and assembly of mitochondrial complex I and IV. The mitochondrial interactomes of wild type and mutant IDE are highly similar and do not reveal any likely proteolytic IDE substrates. We speculate that IDE could adopt similar additional non-proteolytic functions in mitochondria as in the cytosol, acting as a chaperone and contributing to protein homeostasis and stress responses.

Assembly of mitochondrial complex I

Assembly of mitochondrial complex I

Pleiotropic roles of evolutionarily conserved signaling intermediate in toll pathway (ECSIT) in pathophysiology.

The evolutionarily conserved signaling intermediate in toll pathway (ECSIT) is a cytosolic adaptor protein associated with the toll-like receptor pathway. It has a distinct N-terminal mitochondrial targeting sequence, pentatricopeptide repeat motif, and a C-terminal pleckstrin homology domain. ECSIT regulates many biological processes like embryonic development, inflammation, cardiac function, and assembly of mitochondrial complex I. Besides, ECSIT also interacts with multiple signaling intermediates like tumor necrosis receptor associated factor 6and retinoic acid inducible gene 1as well as regulates various pathways in the microcellular environment. However, molecular details of ECSIT functions in pathophysiology remain elusive. This review summarizes the diverse functions of ECSIT and its involvement in pathophysiological conditions such as Alzheimer's, oxidative stress, and infection.

Inhibition of DRP1 Impedes Zygotic Genome Activation and Preimplantation Development in Mice.

Mitochondrion plays an indispensable role during preimplantation embryo development. Dynamic-related protein 1 (DRP1) is critical for mitochondrial fission and controls oocyte maturation. However, its role in preimplantation embryo development is still lacking. In this study, we demonstrate that inhibition of DRP1 activity by mitochondrial division inhibitor-1, a small molecule reported to specifically inhibit DRP1 activity, can cause severe developmental arrest of preimplantation embryos in a dose-dependent manner in mice. Meanwhile, DRP1 inhibition resulted in mitochondrial dysfunction including decreased mitochondrial activity, loss of mitochondrial membrane potential, reduced mitochondrial copy number and inadequate ATP by disrupting both expression and activity of DRP1 and mitochondrial complex assembly, leading to excessive ROS production, severe DNA damage and cell cycle arrest at 2-cell embryo stage. Furthermore, reduced transcriptional and translational activity and altered histone modifications in DRP1-inhibited embryos contributed to impeded zygotic genome activation, which prevented early embryos from efficient development beyond 2-cell embryo stage. These results show that DRP1 inhibition has potential cytotoxic effects on mammalian reproduction, and DRP1 inhibitor should be used with caution when it is applied to treat diseases. Additionally, this study improves our understanding of the crosstalk between mitochondrial metabolism and zygotic genome activation.

Optic atrophy–associated TMEM126A is an assembly factor for the ND4-module of mitochondrial complex I

Mitochondrial disease is a debilitating condition with a diverse genetic etiology. Here, we report that TMEM126A, a protein that is mutated in patients with autosomal-recessive optic atrophy, participates directly in the assembly of mitochondrial complex I. Using a combination of genome editing, interaction studies, and quantitative proteomics, we find that loss of TMEM126A results in an isolated complex I deficiency and that TMEM126A interacts with a number of complex I subunits and assembly factors. Pulse-labeling interaction studies reveal that TMEM126A associates with the newly synthesized mitochondrial DNA (mtDNA)-encoded ND4 subunit of complex I. Our findings indicate that TMEM126A is involved in the assembly of the ND4 distal membrane module of complex I. In addition, we find that the function of TMEM126A is distinct from its paralogue TMEM126B, which acts in assembly of the ND2-module of complex I.

Parkin expression reverses mitochondrial dysfunction in fused in sarcoma-induced amyotrophic lateral sclerosis.

Fused in sarcoma (FUS) is a DNA/RNA-binding protein associated with amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration. The exact molecular mechanisms by which FUS results in neurotoxicity have not yet been fully elucidated. Here, we found that parkin is a genetic suppressor of defective phenotypes induced by exogenous human wild type FUS in Drosophila. Although parkin overexpression did not modulate the FUS protein expression level, the locomotive defects in FUS-expressing larvae and adult flies were rescued by parkin expression. We found that FUS expression in muscle tissues resulted in a reduction of the levels and assembly of mitochondrial complex I and III subunits, as well as decreased ATP. Remarkably, expression of parkin suppressed these mitochondrial dysfunctions. Our results indicate parkin as a neuroprotective regulator of FUS-induced proteinopathy by recovering the protein levels of mitochondrial complexes I and III. Our findings on parkin-mediated neuroprotection may expand our understanding of FUS-induced ALS pathogenesis.

EMP18 functions in mitochondrial atp6 and cox2 transcript editing and is essential to seed development in maize.

RNA editing plays an important role in organellar gene expression in plants, and pentatricopeptide repeat (PPR) proteins are involved in this function. Because of its large family size, many PPR proteins are not known for their function and roles in plant growth and development. Through genetic and molecular analyses of the empty pericarp18 (emp18) mutant in maize (Zea mays), we cloned the Emp18 gene, revealed its molecular function, and defined its role in the mitochondrial complex assembly and seed development. Emp18 encodes a mitochondrial-localized DYW-PPR protein. Null mutation of Emp18 arrests embryo and endosperm development at an early stage in maize, resulting in embryo lethality. Mutants are deficient in the cytidine (C)-to-uridine (U) editing at atp6-635 and cox2-449, which converts a Leu to Pro in ATP6 and a Met to Thr in Cox2. The atp6 gene encodes the subunit a of F1 Fo -ATPase. The Leu to Pro alteration disrupts an α-helix of subunit a, resulting in a dramatic reduction in assembly and activity of F1 Fo -ATPase holoenzyme and an accumulation of free F1 -subcomplex. These results demonstrate that EMP18 functions in the C-to-U editing of atp6 and cox2, and is essential to mitochondrial biogenesis and seed development in maize.

Alterations of mitochondrial protein assembly and jasmonic acid biosynthesis pathway in Honglian (HL)-type cytoplasmic male sterility rice.

It has been suggested that the mitochondrial chimeric gene orfH79 is the cause for abortion of microspores in Honglian cytoplasmic male sterile rice, yet little is known regarding its mechanism of action. In this study, we used a mass spectrometry-based quantitative proteomics strategy to compare the mitochondrial proteome between the sterile line Yuetai A and its fertile near-isogenic line Yuetai B. We discovered a reduced quantity of specific proteins in mitochondrial complexes in Yuetai A compared with Yuetai B, indicating a defect in mitochondrial complex assembly in the sterile line. Western blotting showed that ORFH79 protein and ATP1 protein, an F(1) sector component of complex V, are both associated with large protein complexes of similar size. Respiratory complex activity assays and transmission electron microscopy revealed functional and morphological defects in the mitochondria of Yuetai A when compared with Yuetai B. In addition, we identified one sex determination TASSELSEED2-like protein increased in Yuetai A, leading to the discovery of an aberrant variation of the jasmonic acid pathway during the development of microspores.

The role of Ndufaf2 in the assembly of mitochondrial complex I

The role of Ndufaf2 in the assembly of mitochondrial complex I

Melatonin improves mitochondrial respiratory chain activity and liver morphology in ob/ob mice

In previous studies, we have shown that mitochondrial respiratory chain (MRC) activity is decreased in patients with nonalcoholic steatohepatitis and in ob/ob mice and that peroxynitrite plays a pathogenic role. The present study examined whether melatonin, a peroxynitrite scavenger, prevents: (i) the in vitro effects of peroxynitrite on normal mitochondrial proteins and (ii) the development of nonalcoholic liver disease, MRC dysfunction and proteomic changes found in the mitochondrial complexes from ob/ob mice. We studied MRC activity, assembly of mitochondrial complexes and its subunits in normal mitochondrial proteins exposed to peroxynitrite in the absence and presence of melatonin. The same studies were done in mitochondrial proteins from ob/ob mice untreated and treated with melatonin. Preincubation of mitochondrial proteins from wild-type mice with melatonin prevented 3-tyrosine nitration of these proteins, eliminated the reduction in the MRC activity, the defect in the assembly of mitochondrial complexes and degradation of their subunits induced by peroxynitrite in vitro. Moreover, treatment of ob/ob mice with 10 mg/kg/day melatonin for 12 wk reduced oxidative and nitrosative stress, prevented the loss of MRC activity, protected their complexes and subunits from degradation, and favored assembling of mitochondrial complexes. In addition, this treatment improved fatty liver, decreased hepatic triglyceride concentration and increased apolipoprotein B100 in liver tissue. In conclusion, melatonin prevents the effects of peroxynitrite on mitochondrial proteins in vitro and administration of melatonin to ob/ob mice normalizes liver morphology, mitochondrial dysfunction and assembly of MRC complexes.

Cytosolic signaling protein Ecsit also localizes to mitochondria where it interacts with chaperone NDUFAF1 and functions in complex I assembly

Ecsit is a cytosolic adaptor protein essential for inflammatory response and embryonic development via the Toll-like and BMP (bone morphogenetic protein) signal transduction pathways, respectively. Here, we demonstrate a mitochondrial function for Ecsit (an evolutionary conserved signaling intermediate in Toll pathways) in the assembly of mitochondrial complex I (NADH:ubiquinone oxidoreductase). An N-terminal targeting signal directs Ecsit to mitochondria, where it interacts with assembly chaperone NDUFAF1 in 500- to 850-kDa complexes as demonstrated by affinity purification and vice versa RNA interference (RNAi) knockdowns. In addition, Ecsit knockdown results in severely impaired complex I assembly and disturbed mitochondrial function. These findings support a function for Ecsit in the assembly or stability of mitochondrial complex I, possibly linking assembly of oxidative phosphorylation complexes to inflammatory response and embryonic development.

Rapid rates of newly synthesized mitochondrial protein degradation are significantly affected by the generation of mitochondrial free radicals

Exposure of biological material to high levels of free radicals causes extensive cellular damage. Reactive oxygen species (ROS) generated by mitochondria have been associated with a variety of diseases and aging. We investigated the effect of low-level mitochondrial ROS production on newly synthesized mitochondrial proteins which are potentially vulnerable to mitochondrial ROS due to their location and unfolded state. We show that elevated mitochondrial ROS increases the degradation of newly synthesized mitochondrial proteins with some proteins more sensitive than others. In the long term reduced assembly of mitochondrial complexes would affect mitochondrial function and may trigger a vicious cycle of mitochondrial ROS production.