Mitochondrial Oxidative Phosphorylation Disorders Research Articles

Oxidative Phosphorylation Dysfunction Modifies the Cell Secretome.

Mitochondrial oxidative phosphorylation disorders are extremely heterogeneous conditions. Their clinical and genetic variability makes the identification of reliable and specific biomarkers very challenging. Until now, only a few studies have focused on the effect of a defective oxidative phosphorylation functioning on the cell’s secretome, although it could be a promising approach for the identification and pre-selection of potential circulating biomarkers for mitochondrial diseases. Here, we review the insights obtained from secretome studies with regard to oxidative phosphorylation dysfunction, and the biomarkers that appear, so far, to be promising to identify mitochondrial diseases. We propose two new biomarkers to be taken into account in future diagnostic trials.

Mitochondrial oxidative phosphorylation disorders in children: Phenotypic, genotypic and biochemical correlations in 85 patients from South India

Mitochondrial oxidative phosphorylation (OXPHOS) disorders account for a variety of neuromuscular disorders in children. In this study mitochondrial respiratory chain enzymes were assayed in muscle tissue in a large cohort of children with varied neuromuscular presentations from June 2011 to December 2013. The biochemical enzyme deficiencies were correlated with the phenotypes, magnetic resonance imaging, histopathology and genetic findings to reach a final diagnosis. There were 85 children (mean age: 6.9±4.7years, M:F:2:1) with respiratory chain enzyme deficiency which included: isolated complex I (n=50, 60%), multiple complexes (n=24, 27%), complex IV (n=8, 9%) and complex III deficiencies (n=3, 4%). The most common neurological findings were ataxia (59%), hypotonia (59%) and involuntary movements (49%). A known mitochondrial syndrome was diagnosed in 27 (29%) and non-syndromic presentations in 57 (71%). Genetic analysis included complete sequencing of mitochondrial genome, SURF1, POLG1&2. It revealed variations in mitochondrial DNA (n=8), SURF1 (n=5), and POLG1 (n=3). This study, the first of its kind from India, highlights the wide range of clinical and imaging phenotypes and genetic heterogeneity in children with mitochondrial oxidative phosphorylation disorders.

Proteomics and mitochondrial disease

Disorders of mitochondrial oxidative phosphorylation (OXPHOS) present with almost any symptom, at any age with any mode of inheritance. OXPHOS disorders are the most common group of inherited metabolic diseases and occur in ~1/5000 births. There is no ‘gold standard’ diagnostic test and most paediatric diagnoses rely on finding an enzyme defect while molecular investigations are typically restricted to testing for common mutations or sequencing of specific candidate genes. While next generation sequencing (NGS) approaches have yielded great insights into the identification of pathogenic mutations, these generally focus on sequencing of known mitochondrial and disease related genes, yielding a molecular diagnosis in ~60% of cases. Recent advances in proteomics have improved such that much of the human cellular proteome can now be determined in a matter of hours. In addition, with the use of quantitaive approaches including stable isotope labeling of cells in culture (SILAC), differences between the proteomes of diseased and controls cells can be measured. We are now investigating the use of quantitative proteomics as a complementery tool to NGS in the molecular diagnosis of OXPHOS disease, and to ultimately identify novel candidate genes that cause disease.

Normal oxidative phosphorylation in intestinal smooth muscle of childhood chronic intestinal pseudo-obstruction

Chronic intestinal pseudo-obstruction (CIPO) is a severe disease of the digestive tract motility. In pediatric population, CIPO remains of unknown origin for most patients. Chronic intestinal pseudo-obstruction is also a common feature in the course of mitochondrial oxidative phosphorylation disorders related for some patients to mutations in TYMP, POLG1, mtDNA tRNA(leu(UUR)) or tRNA(lys) genes. We hypothesized that CIPOs could be the presenting symptom of respiratory chain enzyme deficiency and thus we investigated oxidative phosphorylation in small bowel and/or colon smooth muscle of primary CIPO children. We studied eight children with CIPO and 12 pediatric controls. We collected clinical, radiological and pathological data and measured respiratory chain enzymatic activity in isolated smooth muscle of the small bowel and/or the colon. We also sequenced TYMP, POLG, mtDNA tRNA(leu(UUR)) and tRNA(lys) genes. Neither pathological nor radiological data were in favor of a mitochondrial dysfunction. No respiratory chain enzyme deficiency was detected in CIPO children. In myogenic CIPO, respiratory enzymes and citrate synthase activities were increased in small bowel and/or colon whereas no abnormality was noted in neurogenic and unclassified CIPO. Levels of enzyme activities were higher in control small bowel than in control colon muscle. Sequencing of TYMP, POLG, mtDNA tRNA(leu(UUR)) and tRNA(lys) genes and POLG gene did not reveal mutation for any of the patients. The normal enzymatic activities as the lack of radiological and genetic abnormalities indicate that, at variance with adult patients, oxidative phosphorylation deficiency is not a common cause of childhood CIPO.

Mitochondrial Oxidative Phosphorylation Disorders Presenting in Neonates: Clinical Manifestations and Enzymatic and Molecular Diagnoses

Mitochondrial Oxidative Phosphorylation Disorders Presenting in Neonates: Clinical Manifestations and Enzymatic and Molecular Diagnoses

Mitochondrial Disease in Autism Spectrum Disorder Patients: A Cohort Analysis

BackgroundPrevious reports indicate an association between autism spectrum disorders (ASD) and disorders of mitochondrial oxidative phosphorylation. One study suggested that children with both diagnoses are clinically indistinguishable from children with idiopathic autism. There are, however, no detailed analyses of the clinical and laboratory findings in a large cohort of these children. Therefore, we undertook a comprehensive review of patients with ASD and a mitochondrial disorder.Methodology/Principal FindingsWe reviewed medical records of 25 patients with a primary diagnosis of ASD by DSM-IV-TR criteria, later determined to have enzyme- or mutation-defined mitochondrial electron transport chain (ETC) dysfunction. Twenty-four of 25 patients had one or more major clinical abnormalities uncommon in idiopathic autism. Twenty-one patients had histories of significant non-neurological medical problems. Nineteen patients exhibited constitutional symptoms, especially excessive fatigability. Fifteen patients had abnormal neurological findings. Unusual developmental phenotypes included marked delay in early gross motor milestones (32%) and unusual patterns of regression (40%). Levels of blood lactate, plasma alanine, and serum ALT and/or AST were increased at least once in 76%, 36%, and 52% of patients, respectively. The most common ETC disorders were deficiencies of complex I (64%) and complex III (20%). Two patients had rare mtDNA mutations of likely pathogenicity.Conclusions/SignificanceAlthough all patients' initial diagnosis was idiopathic autism, careful clinical and biochemical assessment identified clinical findings that differentiated them from children with idiopathic autism. These and prior data suggest a disturbance of mitochondrial energy production as an underlying pathophysiological mechanism in a subset of individuals with autism.

Mitochondrial Oxidative Phosphorylation Disorders Presenting in Neonates: Clinical Manifestations and Enzymatic and Molecular Diagnoses

The goals were to examine the frequency of perinatal manifestations of mitochondrial oxidative phosphorylation disorders within a population-based cohort, to characterize these manifestations, to identify a possible association between these manifestations and diagnoses at a later age, and to identify possible associations between perinatal complications and specific disorders. We conducted a retrospective review of clinical and laboratory records for all patients with definitive oxidative phosphorylation disorders who were diagnosed and treated at the Royal Children's Hospital in Melbourne between 1975 and 2006 (N = 107; male/female ratio: 1.41). Neonatal presentation was recorded for 32 of 107 patients (male/female ratio: 1:1), including 19 who presented on day 1 of life. Prematurity (gestational age of <37 weeks) was noted for 12.6% of the 107 patients. Of the 85 infants with known birth weights, 24 were in the <or=10th percentile for gestational age (11 with complex I deficiency), and 9 of those (6 with complex I deficiency) were in the <3rd percentile. The most common presenting neonatal symptoms after the first day of life were poor feeding, recurrent vomiting, and failure to thrive. We noted 3 main clinical neonatal forms of oxidative phosphorylation disorders (encephalomyopathic, hepatointestinal, and cardiac). Of the 32 infants, 28 died (13 in the neonatal period). Complex I deficiency was identified for 15 neonates, combined complexes I, III, and IV deficiency for 7 neonates, and combined complexes I and IV deficiency for 3 neonates. No neonates had complex IV deficiency. Six neonates had nuclear mutations, and 2 neonates had the mitochondrial DNA 8993T>G mutation. Oxidative phosphorylation disorders present commonly in the neonatal period. The combination of nonspecific manifestations such as prematurity and intrauterine growth retardation with early postnatal decompensation or poor feeding or vomiting and persistent lactic acidosis should suggest the possibility of an oxidative phosphorylation disorder.

Approaches to Finding the Molecular Basis of Mitochondrial Oxidative Phosphorylation Disorders

Inherited disorders of mitochondrial oxidative phosphorylation are the most common group of inborn errors of metabolism and cause a wide range of clinical presentations. Mitochondrial DNA encodes 13 protein subunits required for oxidative phosphorylation plus 22 transfer RNAs and two ribosomal RNAs, and mutations in most of these genes cause human disease. Nuclear genes encode most of the protein subunits and all other proteins required for mitochondrial biogenesis and mitochondrial DNA replication and expression. Mutations in 64 nuclear genes and 34 mitochondrial genes are now known to cause mitochondrial disease and many novel mitochondrial disease genes await discovery. The genetic complexity of oxidative phosphorylation means that maternal, autosomal recessive, autosomal dominant and X-linked modes of inheritance can occur, along with de novo mutations. This complexity presents a challenge in planning efficient molecular genetic diagnosis of patients with suspected mitochondrial disease. In some situations, clinical phenotype can be strongly predictive of the underlying genotype. However, more often this is not the case and it is usually helpful, particularly with pediatric patients, to determine whether the activity of one or more of the individual oxidative phosphorylation enzymes is deficient before proceeding with mutation analysis. In this review we will summarize the genetic bases of mitochondrial disease and discuss some approaches to integrate information from clinical presentation, laboratory findings, family history, and imaging to guide molecular investigation.

Reconstructing the evolution of the mitochondrial ribosomal proteome

For production of proteins that are encoded by the mitochondrial genome, mitochondria rely on their own mitochondrial translation system, with the mitoribosome as its central component. Using extensive homology searches, we have reconstructed the evolutionary history of the mitoribosomal proteome that is encoded by a diverse subset of eukaryotic genomes, revealing an ancestral ribosome of alpha-proteobacterial descent that more than doubled its protein content in most eukaryotic lineages. We observe large variations in the protein content of mitoribosomes between different eukaryotes, with mammalian mitoribosomes sharing only 74 and 43% of its proteins with yeast and Leishmania mitoribosomes, respectively. We detected many previously unidentified mitochondrial ribosomal proteins (MRPs) and found that several have increased in size compared to their bacterial ancestral counterparts by addition of functional domains. Several new MRPs have originated via duplication of existing MRPs as well as by recruitment from outside of the mitoribosomal proteome. Using sensitive profile–profile homology searches, we found hitherto undetected homology between bacterial and eukaryotic ribosomal proteins, as well as between fungal and mammalian ribosomal proteins, detecting two novel human MRPs. These newly detected MRPs constitute, along with evolutionary conserved MRPs, excellent new screening targets for human patients with unresolved mitochondrial oxidative phosphorylation disorders.

Evaluation of mitochondrial membrane potential using a computerized device with a tetraphenylphosphonium-selective electrode

Mitochondrial membrane potential (Δψ m) plays important roles in the normal function of cells and in pathobiochemical situations. The application of ion-selective electrodes for the measurement of Δψ m is important for studying normal biological reactions and pathways and mitochondrial diseases. We constructed and optimized a computerized device for real-time monitoring of the Δψ m, which included modification of tetraphenylphosphonium (TPP +)-selective membrane that improved reproducibility of the TPP +-selective electrode. Application of MATLAB software increased the sensitivity of the system. We tested our improved device for membrane potential measurements of isolated mitochondria (in absolute scale of millivolts). In addition, we assessed relative changes of Δψ m (as changes in TPP + concentration) of digitonin-permeabilized cells (hepatocytes, control transmitochondrial cybrids, HeLa G and BSC-40) after addition of substrates, inhibitors, and uncoupler of respiratory chain. Our system can be successfully used for studies of many aspects of the regulation of mitochondrial bioenergetics and as a diagnostic tool for mitochondrial oxidative phosphorylation disorders.

Mitochondrial disorders: prevalence, myths and advances.

Disorders of mitochondrial oxidative phosphorylation (OXPHOS) are renowned for their variability in clinical features and genetic causes. This makes it difficult to determine their true prevalence, but recent studies have documented a minimum birth prevalence of 13.1/100000 or 1/7634 for oxidative phosphorylation disorders with onset at any age. This clearly remains an underestimate but it indicates that oxidative phosphorylation disorders can be regarded as the most common group of inborn errors of metabolism. Pathogenic mutations causing human oxidative phosphorylation disorders have now been identified in more than 30 of the 37 mitochondrial DNA genes and in more than 30 nuclear genes. Most of the nuclear gene defects cause autosomal recessive diseases, but autosomal dominant and X-linked disorders also occur. It is likely that at least another 30, and perhaps over 100, nuclear-encoded oxidative phosphorylation disorders await identification. Oxidative phosphorylation genetics are complex and there appear to be a number of common misconceptions about mitochondrial DNA mutations that may impede optimal investigation and management of patients. In our experience, mitochondrial DNA mutations are not a negligible cause of OXPHOS disorders in children but account for 20-25% of cases. Similarly, a family history suggesting maternal inheritance is the exception rather than the norm for children with mitochondrial DNA mutations, many of whom have de novo mutations. Only some mitochondrial DNA mutations disappear from cultured cells, so deficient enzyme activity in fibroblasts does not imply the presence of a nuclear defect. Finally, it is still widely thought that there are very few reproductive options that can be offered to women at risk of transmitting a mitochondrial DNA mutation. While a cautious approach is needed, there is now a consensus that prenatal diagnosis should be offered to some women, particularly those at lower recurrence risk. Preimplantation genetic diagnosis can also be an option.

Screening Human EST Database for Identification of Candidate Genes in Respiratory Chain Deficiency

Disorders of mitochondrial oxidative phosphorylation (OXPHOS) are now recognized as major causes of human metabolic diseases and several mutations of mitochondrial and nuclear genes encoding respiratory chain components have been reported. Interestingly, mutations of nuclear genes involved in mitochondrial respiratory chain assembly, protein trafficking, and iron metabolism are also known to alter oxidative phosphorylation. While several hundred of these genes have been described in yeast, only a few nuclear genes have been hitherto identified in humans. Yeast gene databases present therefore an invaluable tool for identification of human homologues that should be regarded as candidate genes in OXPHOS diseases. In an attempt to identify the human counterparts of yeast genes, we developed a systematic comparison of yeast protein sequences to the GenBank dbEST database. Starting from 340 yeast protein sequences as templates, we searched the human dbEST counterparts using the BLAST similarity searching program and identified 102 groups of human EST likely to represent orthologues of yeast genes because of significant homology. This collection of human genes possibly related to mitochondrial OXPHOS may help identify nuclear genes responsible of mitochondrial disorders.