Correcting a pathogenic mitochondrial DNA mutation by base editing in mice.

Primary mitochondrial disorders are most often caused by deleterious mutations in the mitochondrial DNA (mtDNA). Here, we used a mitochondrial DddA-derived cytosine base editor (DdCBE) to introduce a compensatory edit in a mouse model that carries the pathological mutation in the mitochondrial transfer RNA (tRNA) alanine (mt-tRNA) gene. Because the original m.

Mechanisms and pathologies of human mitochondrial DNA replication and deletion formation.

Human mitochondria possess a multi-copy circular genome, mitochondrial DNA (mtDNA), that is essential for cellular energy metabolism. The number of copies of mtDNA per cell, and their integrity, are maintained by nuclear-encoded mtDNA replication and repair machineries. Aberrant mtDNA replication and mtDNA breakage are believed to cause deletions within mtDNA.

mitoTALEN reduces the mutant mtDNA load in neurons.

Mutations within mtDNA frequently give rise to severe encephalopathies. Given that a majority of these mtDNA defects exist in a heteroplasmic state, we harnessed the precision of mitochondrial-targeted TALEN (mitoTALEN) to selectively eliminate mutant mtDNA within the CNS of a murine model harboring a heteroplasmic mutation in the mitochondrial tRNA alanine gene (m.5024C>T).

High-throughput single-cell analysis reveals progressive mitochondrial DNA mosaicism throughout life.

Heteroplasmic mitochondrial DNA (mtDNA) mutations are a major cause of inherited disease and contribute to common late-onset human disorders. The late onset and clinical progression of mtDNA-associated disease is thought to be due to changing heteroplasmy levels, but it is not known how and when this occurs. Performing high-throughput single-cell genotyping in two mouse models of human mtDNA disease, we saw unanticipated cell-to-cell differences in mtDNA heteroplasmy levels that emerged prenatally and progressively increased throughout life.

Cultured lymphocytes' mitochondrial genome integrity is not altered by cladribine.

Cladribine tablets are a treatment for multiple sclerosis with effects on lymphocytes, yet its mode of action has not been fully established. Here, we analyzed the effects of cladribine on mitochondrial DNA integrity in lymphocytes. We treated cultured human T-cell lines (CCRF-CEM and Jurkat) with varying concentrations of cladribine to mimic the slow cell depletion observed in treated patients.

Cell lineage-specific mitochondrial resilience during mammalian organogenesis.

Mitochondrial activity differs markedly between organs, but it is not known how and when this arises. Here we show that cell lineage-specific expression profiles involving essential mitochondrial genes emerge at an early stage in mouse development, including tissue-specific isoforms present before organ formation. However, the nuclear transcriptional signatures were not independent of organelle function.

Large dataset of octocoral mitochondrial genomes provides new insights into mt-mutS evolution and function.

All studied octocoral mitochondrial genomes (mt-genomes) contain a homologue of the Escherichia coli mutS gene, a member of a gene family encoding proteins involved in DNA mismatch repair, other types of DNA repair, meiotic recombination, and other functions. Although mutS homologues are found in all domains of life, as well as viruses, octocoral mt-mutS is the only such gene found in an organellar genome. While the function of mtMutS is not known, its domain architecture, conserved sequence, and presence of several characteristic residues suggest its involvement in mitochondrial DNA repair.

Mitochondrial DNA heteroplasmy is modulated during oocyte development propagating mutation transmission.

Heteroplasmic mitochondrial DNA (mtDNA) mutations are a common cause of inherited disease, but a few recurrent mutations account for the vast majority of new families. The reasons for this are not known. We studied heteroplasmic mice transmitting m.

Accurate mapping of mitochondrial DNA deletions and duplications using deep sequencing.

Deletions and duplications in mitochondrial DNA (mtDNA) cause mitochondrial disease and accumulate in conditions such as cancer and age-related disorders, but validated high-throughput methodology that can readily detect and discriminate between these two types of events is lacking. Here we establish a computational method, MitoSAlt, for accurate identification, quantification and visualization of mtDNA deletions and duplications from genomic sequencing data. Our method was tested on simulated sequencing reads and human patient samples with single deletions and duplications to verify its accuracy.

High-Throughput Detection of mtDNA Mutations Leading to tRNA Processing Errors.

Some mutations in the tRNA genes of mitochondrial DNA (mtDNA) have been demonstrated to affect the processing of the mitochondrial transcriptome in human patients with mitochondrial disease. A recent analysis of mtDNA mutations in 527 human tumors revealed that approximately a quarter of the somatic mt-tRNA gene mutations lead to aberrant processing of the mitochondrial transcriptome in these tumors. Here, we describe a method, based on mtDNA mutations induced by the mtDNA mutator mouse, to map the sites that lead to transcript processing abnormalities.

Characterization of E-cigarette coil temperature and toxic metal analysis by infrared temperature sensing and scanning electron microscopy - energy-dispersive X-ray.

Introduction: Electronic cigarettes (e-cigarettes) have rapidly evolved since their introduction to the U.S. market.

Current progress with mammalian models of mitochondrial DNA disease.

Mitochondrial disorders make up a large class of heritable diseases that cause a broad array of different human pathologies. They can affect many different organ systems, or display very specific tissue presentation, and can lead to illness either in childhood or later in life. While the over 1200 genes encoded in the nuclear DNA play an important role in human mitochondrial disease, it has been known for over 30 years that mutations of the mitochondria's own small, multicopy DNA chromosome (mtDNA) can lead to heritable human diseases.

Extreme heterogeneity of human mitochondrial DNA from organelles to populations.

Contrary to the long-held view that most humans harbour only identical mitochondrial genomes, deep resequencing has uncovered unanticipated extreme genetic variation within mitochondrial DNA (mtDNA). Most, if not all, humans contain multiple mtDNA genotypes (heteroplasmy); specific patterns of variants accumulate in different tissues, including cancers, over time; and some variants are preferentially passed down or suppressed in the maternal germ line. These findings cast light on the origin and spread of mtDNA mutations at multiple scales, from the organelle to the human population, and challenge the conventional view that high percentages of a mutation are required before a new variant has functional consequences.

Mitochondrial stress response triggered by defects in protein synthesis quality control.

Mitochondria have a compartmentalized gene expression system dedicated to the synthesis of membrane proteins essential for oxidative phosphorylation. Responsive quality control mechanisms are needed to ensure that aberrant protein synthesis does not disrupt mitochondrial function. Pathogenic mutations that impede the function of the mitochondrial matrix quality control protease complex composed of AFG3L2 and paraplegin cause a multifaceted clinical syndrome.

Author Correction: MitoTALEN reduces mutant mtDNA load and restores tRNA levels in a mouse model of heteroplasmic mtDNA mutation.

In the version of this article originally published, there was an error in Fig. 1a. The m.

MitoTALEN reduces mutant mtDNA load and restores tRNA levels in a mouse model of heteroplasmic mtDNA mutation.

Mutations in the mitochondrial DNA (mtDNA) are responsible for several metabolic disorders, commonly involving muscle and the central nervous system. Because of the critical role of mtDNA in oxidative phosphorylation, the majority of pathogenic mtDNA mutations are heteroplasmic, co-existing with wild-type molecules. Using a mouse model with a heteroplasmic mtDNA mutation, we tested whether mitochondrial-targeted TALENs (mitoTALENs) could reduce the mutant mtDNA load in muscle and heart.

Genome editing in mitochondria corrects a pathogenic mtDNA mutation in vivo.

Mutations of the mitochondrial genome (mtDNA) underlie a substantial portion of mitochondrial disease burden. These disorders are currently incurable and effectively untreatable, with heterogeneous penetrance, presentation and prognosis. To address the lack of effective treatment for these disorders, we exploited a recently developed mouse model that recapitulates common molecular features of heteroplasmic mtDNA disease in cardiac tissue: the m.

Delivery of mtZFNs into Early Mouse Embryos.

Mitochondrial diseases often result from mutations in the mitochondrial genome (mtDNA). In most cases, mutant mtDNA coexists with wild-type mtDNA, resulting in heteroplasmy. One potential future approach to treat heteroplasmic mtDNA diseases is the specific elimination of pathogenic mtDNA mutations, lowering the level of mutant mtDNA below pathogenic thresholds.

Base-excision repair deficiency alone or combined with increased oxidative stress does not increase mtDNA point mutations in mice.

Mitochondrial DNA (mtDNA) mutations become more prevalent with age and are postulated to contribute to the ageing process. Point mutations of mtDNA have been suggested to originate from two main sources, i.e.

A novel histochemistry assay to assess and quantify focal cytochrome c oxidase deficiency.

Defects in the respiratory chain, interfering with energy production in the cell, are major underlying causes of mitochondrial diseases. In spite of this, the surprising variety of clinical symptoms, disparity between ages of onset, as well as the involvement of mitochondrial impairment in ageing and age-related diseases continue to challenge our understanding of the pathogenic processes. This complexity can be in part attributed to the unique metabolic needs of organs or of various cell types.

Mice lacking the mitochondrial exonuclease MGME1 accumulate mtDNA deletions without developing progeria.

Replication of mammalian mitochondrial DNA (mtDNA) is an essential process that requires high fidelity and control at multiple levels to ensure proper mitochondrial function. Mutations in the mitochondrial genome maintenance exonuclease 1 (MGME1) gene were recently reported in mitochondrial disease patients. Here, to study disease pathophysiology, we generated Mgme1 knockout mice and report that homozygous knockouts develop depletion and multiple deletions of mtDNA.

Increased Total mtDNA Copy Number Cures Male Infertility Despite Unaltered mtDNA Mutation Load.

Mutations of mtDNA cause mitochondrial diseases and are implicated in age-associated diseases and aging. Pathogenic mtDNA mutations are often present in a fraction of all mtDNA copies, and it has been widely debated whether the proportion of mutant genomes or the absolute number of wild-type molecules determines if oxidative phosphorylation (OXPHOS) will be impaired. Here, we have studied the male infertility phenotype of mtDNA mutator mice and demonstrate that decreasing mtDNA copy number worsens mitochondrial aberrations of spermatocytes and spermatids in testes, whereas an increase in mtDNA copy number rescues the fertility phenotype and normalizes testes morphology as well as spermatocyte proteome changes.

A Phenotype-Driven Approach to Generate Mouse Models with Pathogenic mtDNA Mutations Causing Mitochondrial Disease.

Mutations of mtDNA are an important cause of human disease, but few animal models exist. Because mammalian mitochondria cannot be transfected, the development of mice with pathogenic mtDNA mutations has been challenging, and the main strategy has therefore been to introduce mutations found in cell lines into mouse embryos. Here, we describe a phenotype-driven strategy that is based on detecting clonal expansion of pathogenic mtDNA mutations in colonic crypts of founder mice derived from heterozygous mtDNA mutator mice.

Hierarchical RNA Processing Is Required for Mitochondrial Ribosome Assembly.

The regulation of mitochondrial RNA processing and its importance for ribosome biogenesis and energy metabolism are not clear. We generated conditional knockout mice of the endoribonuclease component of the RNase P complex, MRPP3, and report that it is essential for life and that heart and skeletal-muscle-specific knockout leads to severe cardiomyopathy, indicating that its activity is non-redundant. Transcriptome-wide parallel analyses of RNA ends (PARE) and RNA-seq enabled us to identify that in vivo 5' tRNA cleavage precedes 3' tRNA processing, and this is required for the correct biogenesis of the mitochondrial ribosomal subunits.

Tissue-specific modulation of mitochondrial DNA segregation by a defect in mitochondrial division.

Mitochondria are dynamic organelles that divide and fuse by remodeling an outer and inner membrane in response to developmental, physiological and stress stimuli. These events are coordinated by conserved dynamin-related GTPases. The dynamics of mitochondrial morphology require coordination with mitochondrial DNA (mtDNA) to ensure faithful genome transmission, however, this process remains poorly understood.