Mitochondrial Protein Synthesis Research Articles

Integrated Stress Response Couples Mitochondrial Protein Translation With Oxidative Stress Control.

The integrated stress response (ISR) is an evolutionarily conserved process to cope with intracellular and extracellular disturbances. Myocardial infarction is a leading cause of death worldwide. Coronary artery reperfusion, the most effective means to mitigate cardiac damage of myocardial infarction, causes additional reperfusion injury. This study aimed to investigate the role of the ISR in myocardial ischemia/reperfusion (I/R). Cardiac-specific gain- and loss-of-function approaches for the ISR were used in vivo. Myocardial I/R was achieved by ligation of the cardiac left anterior descending artery for 45 minutes followed by reperfusion for different times. Cardiac function was assessed by echocardiography. Cultured H9c2 cells, primary rat cardiomyocytes, and mouse embryonic fibroblasts were used to dissect underlying molecular mechanisms. Tandem mass tag labeling and mass spectrometry was conducted to identify protein targets of the ISR. Pharmacologic means were tested to manipulate the ISR for therapeutic exploration. We show that the PERK (PKR-like endoplasmic reticulum resident kinase)/eIF2α (α subunit of eukaryotic initiation factor 2) axis of the ISR is strongly induced by I/R in cardiomyocytes in vitro and in vivo. We further reveal a physiologic role of PERK/eIF2α signaling by showing that acute activation of PERK in the heart confers robust cardioprotection against reperfusion injury. In contrast, cardiac-specific deletion of PERK aggravates cardiac responses to reperfusion. Mechanistically, the ISR directly targets mitochondrial complexes through translational suppression. We identify NDUFAF2 (NADH:ubiquinone oxidoreductase complex assembly factor 2), an assembly factor of mitochondrial complex I, as a selective target of PERK. Overexpression of PERK suppresses the protein expression of NDUFAF2 and PERK inhibition causes an increase of NDUFAF2. Silencing of NDUFAF2 significantly rescues cardiac cell survival from PERK knockdown under I/R. We show that activation of PERK/eIF2α signaling reduces mitochondrial complex-derived reactive oxygen species and improves cardiac cell survival in response to I/R. Moreover, pharmacologic stimulation of the ISR protects the heart against reperfusion damage, even after the restoration of occluded coronary artery, highlighting clinical relevance for myocardial infarction treatment. These results suggest that the ISR improves cell survival and mitigates reperfusion damage by selectively suppressing mitochondrial protein synthesis and reducing oxidative stress in the heart.

Anti-tumor activity of tigecycline: a review

Tigecycline is a novel glycylcycline antibacterial drug, which shows both antibiotic function and anti-tumor activity. This review summarizes the single and combined use of tigecycline for tumor treatment and the underpinning mechanisms. As an inhibitor for mitochondrial DNA translation, tigecycline affects the proliferation, migration, and invasion of tumor cells mainly through inhibiting mitochondrial protein synthesis and inducing mitochondrial dysfunction. Although the effect of tigecycline monotherapy is controversial, the efficacy of combined use of tigecycline is satisfactory. Therefore, it is important to explore the molecular mechanisms underpinning the anti-tumor activity of tigecycline, with the aim to use it as a cheap and effective new anti-tumor drug.

Assessment of Protein Synthesis in Mitochondria Isolated from Rosette Leaves and Liquid Culture Seedlings of Arabidopsis.

Mitochondria are subcellular organelles with their own genome and expression system, including translation machinery to make proteins. Several independent studies have shown that translation is an essential regulatory step in expression of the plant mitochondrial genome. Thus, the study of mitochondrial translation seems to be crucial for the comprehension of plant mitochondrial biogenesis and maintenance. In organello protein synthesis in isolated mitochondria is a direct method to visualize the translational products of this organellar genetic system. In this method, highly purified, functional mitochondria synthesize proteins in the presence of radiolabeled amino acids, such as methionine, and an energy regeneration system. The labeled, newly synthesized polypeptides are separated by SDS-polyacrylamide gel electrophoresis and are detected by autoradiography. Here we describe the detailed protocol for in organello labeling of translation products that was optimized for mitochondria isolated from rosette leaves and liquid culture seedlings of Arabidopsis thaliana plants.

AMP deamination is sufficient to replicate an atrophy-like metabolic phenotype in skeletal muscle

BackgroundSkeletal muscle atrophy, whether caused by chronic disease, acute critical illness, disuse or aging, is characterized by tissue-specific decrease in oxidative capacity and broad alterations in metabolism that contribute to functional decline. However, the underlying mechanisms responsible for these metabolic changes are largely unknown. One of the most highly upregulated genes in atrophic muscle is AMP deaminase 3 (AMPD3: AMP → IMP + NH3), which controls the content of intracellular adenine nucleotides (AdN; ATP + ADP + AMP). Given the central role of AdN in signaling mitochondrial gene expression and directly regulating metabolism, we hypothesized that overexpressing AMPD3 in muscle cells would be sufficient to alter their metabolic phenotype similar to that of atrophic muscle. MethodsAMPD3 and GFP (control) were overexpressed in mouse tibialis anterior (TA) muscles via plasmid electroporation and in C2C12 myotubes using adenovirus vectors. TA muscles were excised one week later, and AdN were quantified by UPLC. In myotubes, targeted measures of AdN, AMPK/PGC-1α/mitochondrial protein synthesis rates, unbiased metabolomics, and transcriptomics by RNA sequencing were measured after 24 h of AMPD3 overexpression. Media metabolites were measured as an indicator of net metabolic flux. At 48 h, the AMPK/PGC-1α/mitochondrial protein synthesis rates, and myotube respiratory function/capacity were measured. ResultsTA muscles overexpressing AMPD3 had significantly less ATP than contralateral controls (−25%). In myotubes, increasing AMPD3 expression for 24 h was sufficient to significantly decrease ATP concentrations (−16%), increase IMP, and increase efflux of IMP catabolites into the culture media, without decreasing the ATP/ADP or ATP/AMP ratios. When myotubes were treated with dinitrophenol (mitochondrial uncoupler), AMPD3 overexpression blunted decreases in ATP/ADP and ATP/AMP ratios but exacerbated AdN degradation. As such, pAMPK/AMPK, pACC/ACC, and phosphorylation of AMPK substrates, were unchanged by AMPD3 at this timepoint. AMPD3 significantly altered 191 out of 639 detected intracellular metabolites, but only 30 transcripts, none of which encoded metabolic enzymes. The most altered metabolites were those within purine nucleotide, BCAA, glycolysis, and ceramide metabolic pathways. After 48 h, AMPD3 overexpression significantly reduced pAMPK/AMPK (−24%), phosphorylation of AMPK substrates (−14%), and PGC-1α protein (−22%). Moreover, AMPD3 significantly reduced myotube mitochondrial protein synthesis rates (−55%), basal ATP synthase-dependent (−13%), and maximal uncoupled oxygen consumption (−15%). ConclusionsIncreased expression of AMPD3 significantly decreased mitochondrial protein synthesis rates and broadly altered cellular metabolites in a manner similar to that of atrophic muscle. Importantly, the changes in metabolites occurred prior to reductions in AMPK signaling, gene expression, and mitochondrial protein synthesis, suggesting metabolism is not dependent on reductions in oxidative capacity, but may be consequence of increased AMP deamination. Therefore, AMP deamination in skeletal muscle may be a mechanism that alters the metabolic phenotype of skeletal muscle during atrophy and could be a target to improve muscle function during muscle wasting.

Early resistance training-mediated stimulation of daily muscle protein synthetic responses to higher habitual protein intake in middle-aged adults.

The ingestion of protein potentiates the stimulation of myofibrillar protein synthesis rates after an acute bout of resistance exercise. Protein supplementation (eating above the protein Recommended Dietary Allowance) during resistance training has been shown to maximize lean mass and strength gains in healthy young and older adults. Here, contractile, oxidative, and structural protein synthesis were assessed in skeletal muscle in response to a moderate or higher protein diet during the early adaptive phase of resistance training in middle-aged adults. The stimulation of myofibrillar, mitochondrial or collagen protein synthesis rates during 0-3weeks of resistance training is not further enhanced by a higher protein diet. These results show that moderate protein diets are sufficient to support the skeletal muscle adaptive response during the early phase of a resistance training programme. Protein ingestion augments muscle protein synthesis (MPS) rates acutely after resistance exercise and can offset age-related loss in muscle mass. Skeletal muscle contains a variety of protein pools, such as myofibrillar (contractile), mitochondrial (substrate oxidation), and collagen (structural support) proteins, and the sensitivity to nutrition and exercise seems to be dependent on the major protein fraction studied. However, it is unknown how free-living conditions with high dietary protein density and habitual resistance exercise mediates muscle protein subfraction synthesis. Therefore, we investigated the effect of moderate (MOD: 1.06±0.22gkg-1 day-1 ) or high (HIGH: 1.55±0.25gkg-1 day-1 ) protein intake on daily MPS rates within the myofibrillar (MyoPS), mitochondrial (MitoPS) and collagen (CPS) protein fractions in middle-aged men and women (n=20, 47±1years, BMI 28±1kgm-2 ) during the early phase (0-3weeks) of a dietary counselling-controlled resistance training programme. Participants were loaded with deuterated water, followed by daily maintenance doses throughout the intervention. Muscle biopsies were collected at baseline and after weeks 1, 2 and 3. MyoPS in the HIGH condition remained constant (P=1.000), but MOD decreased over time (P=0.023). MitoPS decreased after 0-3weeks when compared to 0-1week (P=0.010) with no effects of protein intake (P=0.827). A similar decline with no difference between groups (P=0.323) was also observed for CPS (P=0.007). Our results demonstrated that additional protein intake above moderate amounts does not potentiate the stimulation of longer-term MPS responses during the early stage of resistance training adaptations in middle-aged adults.

Extracellular Mitochondrial Components and Effects on Cardiovascular Disease.

Besides being powerhouses of the cell, mitochondria released into extracellular space act as intercellular signaling. Mitochondria and their components mediate cell-to-cell communication in free form or embedded in a carrier. The pathogenesis of cardiovascular disease is complex, which shows close relationship with inflammation and metabolic abnormalities. Since mitochondria sustain optimal function of the heart, extracellular mitochondria are emerging as a key regulator in the development of cardiovascular disease. In this review, we provide recent findings in the presence and forms of mitochondria transfer between cells, as well as the effects of these mitochondria on vascular inflammation and ischemic myocardium. Mitochondrial transplantation is a novel treatment paradigm for patients suffering from acute cardiovascular accident and challenges the traditional methods of mitochondria isolation.

Mitochondrial Reactive Oxygen Species Are Essential for the Development of Psoriatic Inflammation

Psoriasis is a common immune-mediated, chronic, inflammatory skin disease that affects approximately 2–3% of the population worldwide. Although there is increasing evidence regarding the essential roles of the interleukin (IL)-23/IL-17 axis and dendritic cell (DC)-T cell crosstalk in the development of skin inflammation, the contributions of mitochondrial function to psoriasis are unclear. In a mouse model of imiquimod (IMQ)-induced psoriasiform skin inflammation, we found that hematopoietic cell-specific genetic deletion of p32/C1qbp, a regulator of mitochondrial protein synthesis and metabolism, protects mice from IMQ-induced psoriatic inflammation. Additionally, we demonstrate that p32/C1qbp is an important regulator of IMQ-induced DC activation, both in vivo and in vitro. We also found that p32/C1qbp-deficient DCs exhibited impaired production of IL-1β, IL-23, and mitochondrial reactive oxygen species (mtROS) after IMQ stimulation. Because the inhibition of mtROS suppressed IMQ-induced DC activation and psoriatic inflammation, we presume that p32/C1qbp and mtROS can serve as therapeutic targets in psoriasis.

Lack of Increase in Muscle Mitochondrial Protein Synthesis During the Course of Aerobic Exercise and Its Recovery in the Fasting State Irrespective of Obesity.

Acute aerobic exercise induces skeletal muscle mitochondrial gene expression, which in turn can increase muscle mitochondrial protein synthesis. In this regard, the peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α), is a master regulator of mitochondrial biogenesis, and thus mitochondrial protein synthesis. However, PGC-1α expression is impaired in muscle of humans with obesity in response to acute aerobic exercise. Therefore, we sought to determine whether muscle mitochondrial protein synthesis is also impaired under the same conditions in humans with obesity. To this end, we measured mitochondrial and mixed-muscle protein synthesis in skeletal muscle of untrained subjects with (body fat: 34.7 ± 2.3%) and without (body fat: 25.3 ± 3.3%) obesity in a basal period and during a continuous period that included a 45 min cycling exercise (performed at an intensity corresponding to 65% of heart rate reserve) and a 3-h post-exercise recovery. Exercise increased PGC-1α mRNA expression in muscle of subjects without obesity, but not in subjects with obesity. However, muscle mitochondrial protein synthesis did not increase in either subject group. Similarly, mixed-muscle protein synthesis did not increase in either group. Concentrations of plasma amino acids decreased post-exercise in the subjects without obesity, but not in the subjects with obesity. We conclude that neither mitochondrial nor mixed-muscle protein synthesis increase in muscle of humans during the course of a session of aerobic exercise and its recovery period in the fasting state irrespective of obesity.Trial Registration: The study has been registered within ClinicalTrials.gov (NCT01824173).

Photobiomodulation- Yes or No?. following laser gingival depigmentation: A case report

Gingival hyperpigmentation in many esthetic concern patient poses a psychological problem and embarrassment. Among the many procedures used for gingival depigmentation, diode laser is considered as one which is highly accepted by the patient due to the absence of bleeding during and after procedure and being a fast and effective method. Literature have also stated the beneficial effect of low dose laser therapy on wound healing due to its potential to increase mitochondrial function, adenosine triphosphate (ATP), RNA, and protein synthesis which may further increase the cellular metabolism resulting in enhancement in wound healing and acceleration of the inflammatory process. This case report presents the use of diode laser for gingival depigmentation followed by laser photobiomodulation in an attempt to fasten the healing.

A mitochondrial cytidine deaminase is responsible for C to U editing of tRNATrp to decode the UGA codon in Trypanosoma brucei

ABSTRACT In kinetoplastid protists, all mitochondrial tRNAs are encoded in the nucleus and imported from the cytoplasm to maintain organellar translation. This also applies to the tryptophanyl tRNA (tRNATrp) encoded by a single-copy nuclear gene, with a CCA anticodon to read UGG codon used in the cytosolic translation. Yet, in the mitochondrion it is unable to decode the UGA codon specifying tryptophan. Following mitochondrial import of tRNATrp, this problem is solved at the RNA level by a single C34 to U34 editing event that creates the UCA anticodon, recognizing UGA. To identify the enzyme responsible for this critical editing activity, we scrutinized the genome of Trypanosoma brucei for putative cytidine deaminases as the most likely candidates. Using RNAi silencing and poisoned primer extension, we have identified a novel deaminase enzyme, named here TbmCDAT for mitochondrial Cytidine Deaminase Acting on tRNA, which is responsible for this organelle-specific activity in T. brucei. The ablation of TbmCDAT led to the downregulation of mitochondrial protein synthesis, supporting its role in decoding the UGA tryptophan codon.

Exploring the Ability of LARS2 Carboxy-Terminal Domain in Rescuing the MELAS Phenotype.

The m.3243A>G mutation within the mitochondrial mt-tRNALeu(UUR) gene is the most prevalent variant linked to mitochondrial encephalopathy with lactic acidosis and stroke-like episodes (MELAS) syndrome. This pathogenic mutation causes severe impairment of mitochondrial protein synthesis due to alterations of the mutated tRNA, such as reduced aminoacylation and a lack of post-transcriptional modification. In transmitochondrial cybrids, overexpression of human mitochondrial leucyl-tRNA synthetase (LARS2) has proven effective in rescuing the phenotype associated with m.3243A>G substitution. The rescuing activity resides in the carboxy-terminal domain (Cterm) of the enzyme; however, the precise molecular mechanisms underlying this process have not been fully elucidated. To deepen our knowledge on the rescuing mechanisms, we demonstrated the interactions of the Cterm with mutated mt-tRNALeu(UUR) and its precursor in MELAS cybrids. Further, the effect of Cterm expression on mitochondrial functions was evaluated. We found that Cterm ameliorates de novo mitochondrial protein synthesis, whilst it has no effect on mt-tRNALeu(UUR) steady-state levels and aminoacylation. Despite the complete recovery of cell viability and the increase in mitochondrial translation, Cterm-overexpressing cybrids were not able to recover bioenergetic competence. These data suggest that, in our MELAS cell model, the beneficial effect of Cterm may be mediated by factors that are independent of the mitochondrial bioenergetics.

Preferential import of queuosine-modified tRNAs into Trypanosoma brucei mitochondrion is critical for organellar protein synthesis.

Transfer RNAs (tRNAs) are key players in protein synthesis. To be fully active, tRNAs undergo extensive post-transcriptional modifications, including queuosine (Q), a hypermodified 7-deaza-guanosine present in the anticodon of several tRNAs in bacteria and eukarya. Here, molecular and biochemical approaches revealed that in the protozoan parasite Trypanosoma brucei, Q-containing tRNAs have a preference for the U-ending codons for asparagine, aspartate, tyrosine and histidine, analogous to what has been described in other systems. However, since a lack of tRNA genes in T. brucei mitochondria makes it essential to import a complete set from the cytoplasm, we surprisingly found that Q-modified tRNAs are preferentially imported over their unmodified counterparts. In turn, their absence from mitochondria has a pronounced effect on organellar translation and affects function. Although Q modification in T. brucei is globally important for codon selection, it is more so for mitochondrial protein synthesis. These results provide a unique example of the combined regulatory effect of codon usage and wobble modifications on protein synthesis; all driven by tRNA intracellular transport dynamics.

The Role of Mitochondrial Mutations and Chronic Inflammation in Diabetes.

Diabetes mellitus and related disorders significantly contribute to morbidity and mortality worldwide. Despite the advances in the current therapeutic methods, further development of anti-diabetic therapies is necessary. Mitochondrial dysfunction is known to be implicated in diabetes development. Moreover, specific types of mitochondrial diabetes have been discovered, such as MIDD (maternally inherited diabetes and deafness) and DAD (diabetes and Deafness). Hereditary mitochondrial disorders are caused by certain mutations in the mitochondrial DNA (mtDNA), which encodes for a substantial part of mitochondrial proteins and mitochondrial tRNA necessary for mitochondrial protein synthesis. Study of mtDNA mutations is challenging because the pathogenic phenotype associated with such mutations depends on the level of its heteroplasmy (proportion of mtDNA copies carrying the mutation) and can be tissue-specific. Nevertheless, modern sequencing methods have allowed describing and characterizing a number of mtDNA mutations associated with human disorders, and the list is constantly growing. In this review, we provide a list of mtDNA mutations associated with diabetes and related disorders and discuss the mechanisms of their involvement in the pathology development.

Schizosaccharomyces pombe Ppr10 and Mpa1 together mediate mitochondrial translational initiation

Pentatricopeptide repeat (PPR) proteins are a large family of proteins that act primarily at different posttranscriptional steps of organellar gene expression. We have previously found that the Schizosaccharomyces pombe PPR protein mpal10 interacts with mitochondrial translational activator Mpa1, and both are essential for mitochondrial protein synthesis. However, it is unclear how these two proteins function in mitochondrial protein synthesis in S. pombe. In this study, we further investigated the role of Ppr10 and Mpa1 in mitochondrial protein synthesis. Mitochondrial translational initiation requires two initiation factors, Mti2 and Mti3, which bind to the small subunit of the mitochondrial ribosome (mt-SSU) during the formation of the mitochondrial translational initiation complex. Using sucrose gradient sedimentation analysis, we found that disruption of ppr10, mpa1, or the PPR motifs in Ppr10 impairs the association of Mti2 and Mti3 with the mt-SSU, suggesting that both Ppr10 and Mpa1 may be required for the interaction of Mti2 and Mti3 with the mt-SSU during the assembly of mitochondrial translational initiation complex. Loss of Ppr10 perturbs the association of mitochondrially encoded cytochrome b (cob1) and cytochrome c oxidase subunit 1 (cox1) mRNAs with assembled mitochondrial ribosomes. Proteomic analysis revealed that a fraction of Ppr10 and Mpa1 copurified with a subset of mitoribosomal proteins. The PPR motifs of Ppr10 are necessary for its interaction with Mpa1 and that disruption of these PPR motifs impairs mitochondrial protein synthesis. Our results suggest that Ppr10 and Mpa1 function together to mediate mitochondrial translational initiation.

Pathogenic variants in MRPL44 cause infantile cardiomyopathy due to a mitochondrial translation defect

Cardiac dysfunction is a common phenotypic manifestation of primary mitochondrial disease with multiple nuclear and mitochondrial DNA pathogenic variants as a cause, including disorders of mitochondrial translation. To date, five patients have been described with pathogenic variants in MRPL44, encoding the ml44 protein which is part of the large subunit of the mitochondrial ribosome (mitoribosome). Three presented as infants with hypertrophic cardiomyopathy, mild lactic acidosis, and easy fatigue and muscle weakness, whereas two presented in adolescence with myopathy and neurological symptoms.We describe two infants who presented with cardiomyopathy from the neonatal period, failure to thrive, hypoglycemia and in one infant lactic acidosis. A decompensation of the cardiac function in the first year resulted in demise. Exome sequencing identified compound heterozygous variants in the MRPL44 gene including the known pathogenic variant c.467 T > G and two novel pathogenic variants. We document a combined respiratory chain enzyme deficiency with emphasis on complex I and IV, affecting heart muscle tissue more than skeletal muscle or fibroblasts. We show this to be caused by reduced mitochondrial DNA encoded protein synthesis affecting all subunits, and resulting in dysfunction of complex I and IV assembly. The degree of oxidative phosphorylation dysfunction correlated with the impairment of mitochondrial protein synthesis due to different pathogenic variants. These functional studies allow for improved understanding of the pathogenesis of MRPL44-associated mitochondrial disorder.

Mitochondrial mistranslation modulated by metabolic stress causes cardiovascular disease and reduced lifespan.

Changes in the rate and fidelity of mitochondrial protein synthesis impact the metabolic and physiological roles of mitochondria. Here we explored how environmental stress in the form of a high‐fat diet modulates mitochondrial translation and affects lifespan in mutant mice with error‐prone (Mrps12ep / ep ) or hyper‐accurate (Mrps12ha / ha ) mitochondrial ribosomes. Intriguingly, although both mutations are metabolically beneficial in reducing body weight, decreasing circulating insulin and increasing glucose tolerance during a high‐fat diet, they manifest divergent (either deleterious or beneficial) outcomes in a tissue‐specific manner. In two distinct organs that are commonly affected by the metabolic disease, the heart and the liver, Mrps12ep / ep mice were protected against heart defects but sensitive towards lipid accumulation in the liver, activating genes involved in steroid and amino acid metabolism. In contrast, enhanced translational accuracy in Mrps12ha / ha mice protected the liver from a high‐fat diet through activation of liver proliferation programs, but enhanced the development of severe hypertrophic cardiomyopathy and led to reduced lifespan. These findings reflect the complex transcriptional and cell signalling responses that differ between post‐mitotic (heart) and highly proliferative (liver) tissues. We show trade‐offs between the rate and fidelity of mitochondrial protein synthesis dictate tissue‐specific outcomes due to commonly encountered stressful environmental conditions or aging.

Expression and analysis of the SAM-dependent RNA methyltransferase Rsm22 from Saccharomyces cerevisiae.

The Saccharomyces cerevisiae Rsm22 protein (Sc-Rsm22), encoded by the nuclear RSM22 (systematic name YKL155c) gene, is a distant homologue of Rsm22 from Trypanosoma brucei (Tb-Rsm22) and METTL17 from mouse (Mm-METTL17). All three proteins have been shown to be associated with mitochondrial gene expression, and Sc-Rsm22 has been documented to be essential for mitochondrial respiration. The Sc-Rsm22 protein comprises a polypeptide of molecular weight 72.2 kDa that is predicted to harbor an N-terminal mitochondrial targeting sequence. The precise physiological function of Rsm22-family proteins is unknown, and no structural information has been available for Sc-Rsm22 to date. In this study, Sc-Rsm22 was expressed and purified in monomeric and dimeric forms, their folding was confirmed by circular-dichroism analyses and their low-resolution structures were determined using a small-angle X-ray scattering (SAXS) approach. The solution structure of the monomeric form of Sc-Rsm22 revealed an elongated three-domain arrangement, which differs from the shape of Tb-Rsm22 in its complex with the mitochondrial small ribosomal subunit in T. brucei (PDB entry 6sg9). A bioinformatic analysis revealed that the core domain in the middle (Leu117-Asp462 in Sc-Rsm22) resembles the corresponding region in Tb-Rsm22, including a Rossmann-like methyltransferase fold followed by a zinc-finger-like structure. The latter structure is not present in this position in other methyltransferases and is therefore a unique structural motif for this family. The first half of the C-terminal domain is likely to form an OB-fold, which is typically found in RNA-binding proteins and is also seen in the Tb-Rsm22 structure. In contrast, the N-terminal domain of Sc-Rsm22 is predicted to be fully α-helical and shares no sequence similarity with other family members. Functional studies demonstrated that the monomeric variant of Sc-Rsm22 methylates mitochondrial tRNAs in vitro. These data suggest that Sc-Rsm22 is a new and unique member of the RNA methyltransferases that is important for mitochondrial protein synthesis.

Telencephalon-specific Alkbh1 conditional knockout mice display hippocampal atrophy and impaired learning.

AlkB homolog 1 (ALKBH1) is responsible for the biogenesis of 5-formylcytidine (f5 C) on mitochondrial tRNAMet and essential for mitochondrial protein synthesis. The brain, especially the hippocampus, is highly susceptible to mitochondrial dysfunction; hence, the maintenance of mitochondrial activity is strongly required to prevent disorders associated with hippocampal malfunction. To study the role of ALKBH1 in the hippocampus, we generated dorsal telencephalon-specific Alkbh1 conditional knockout (cKO) mice in inbred C57BL/6 background. These mice showed reduced activity of the respiratory chain complex, hippocampal atrophy, and CA1 pyramidal neuron abnormalities. Furthermore, performances in the fear-conditioning and Morris water maze tests in cKO mice indicated that the hippocampal abnormalities led to impaired hippocampus-dependent learning. These findings indicate critical roles of ALKBH1 in the hippocampus.

The Role of Mitochondrial Dysfunction in Vascular Disease, Tumorigenesis, and Diabetes.

Mitochondrial dysfunction is known to be associated with a wide range of human pathologies, such as cancer, metabolic, and cardiovascular diseases. One of the possible ways of mitochondrial involvement in the cellular damage is excessive production of reactive oxygen and nitrogen species (ROS and RNS) that cannot be effectively neutralized by existing antioxidant systems. In mitochondria, ROS and RNS can contribute to protein and mitochondrial DNA (mtDNA) damage causing failure of enzymatic chains and mutations that can impair mitochondrial function. These processes further lead to abnormal cell signaling, premature cell senescence, initiation of inflammation, and apoptosis. Recent studies have identified numerous mtDNA mutations associated with different human pathologies. Some of them result in imbalanced oxidative phosphorylation, while others affect mitochondrial protein synthesis. In this review, we discuss the role of mtDNA mutations in cancer, diabetes, cardiovascular diseases, and atherosclerosis. We provide a list of currently described mtDNA mutations associated with each pathology and discuss the possible future perspective of the research.

Skeletal Muscle Mitochondrial Bioenergetics in a Model of Primary Osteoarthritis after Treatment with Rapamycin and/or Metformin

Impaired mitochondrial function is associated with the loss of skeletal muscle function during aging and age-related osteoarthritis (OA). Emerging evidence suggests Rapamycin (Rap), metformin (Met) or the combination of rapamycin plus metformin (Rap+Met) can extend lifespan and preserve skeletal muscle mass and function during aging. However, the mechanisms of action are not completely understood and have not been studied in specific models of age-related disease. The goal of this study was to test to test the hypothesis that Rap, Met, or Rap+Met would improve skeletal muscle mitochondrial function in an animal model of naturally occurring, age-related OA. We chose the Dunkin Hartley guinea pig because they are an outbred strain that develops naturally occurring OA by 5 months of age with a progression and pathology that highly resembles aging humans. After 5 months of age, Dunkin Hartley guinea pigs also have an age-related decline in skeletal muscle mitochondrial protein synthesis rates. Therefore, 5-month old Dunkin Hartley guinea pigs were fed a standard diet (Con; n=8), or a diet enriched with Rap (14ppm; n=8), Met (1000ppm, n=8) or Rap+Met (14+1000ppm; n=7) for 12-weeks. Mitochondrial respiration and hydrogen peroxide (H2O2) emissions were evaluated in permeabilized muscle fibers from the soleus via high-resolution respirometry and fluorometry using either a saturating bolus or titrating doses of ADP. Michaelis-Menten kinetics was used to evaluate Vmax and ADP sensitivity. All comparisons were made to control group. In the current study, Met had no effect while Rap and Rap+Met modified skeletal muscle mitochondrial bioenergetics only when ADP was titrated but not with a saturating bolus. Specifically, Rap (P<0.05) and Rap+Met (P=0.07) increased ADP sensitivity but decreased (P<0.05) maximal complex-I (CI) linked respiration. Rap and Rap+Met also tended to decrease mitochondrial H2O2 emissions but this trend was no longer apparent when mitochondrial H2O2 emissions were expressed relative to respiration. The decrease in CI-linked respiration and H2O2 emissions was attributed to both a lower CI protein abundance and a decreased index of intrinsic mitochondrial function. This is the first inquiry into how lifespan extending treatments can influence skeletal muscle mitochondria in a model of age-related OA. Further research is needed to understand if increased mitochondrial ADP sensitivity at the cost of CI-linked maximal capacity by Rap and Rap+Met is a beneficial strategy to maintain cellular energy homeostasis during age-related OA.