Mitochondrial Genomic Instability Research Articles

Mitochondrial DNA breaks activate an integrated stress response to reestablish homeostasis

Mitochondrial DNA double-strand breaks (mtDSBs) lead to the degradation of circular genomes and a reduction in copy number; yet, the cellular response in human cells remains elusive. Here, using mitochondrial-targeted restriction enzymes, we show that a subset of cells with mtDSBs exhibited defective mitochondrial protein import, reduced respiratory complexes, and loss of membrane potential. Electron microscopy confirmed the altered mitochondrial membrane and cristae ultrastructure. Intriguingly, mtDSBs triggered the integrated stress response (ISR) via the phosphorylation of eukaryotic translation initiation factor 2α (eIF2α) by DELE1 and heme-regulated eIF2α kinase (HRI). When ISR was inhibited, the cells experienced intensified mitochondrial defects and slower mtDNA recovery post-breakage. Lastly, through proteomics, we identified ATAD3A-a membrane-bound protein interacting with nucleoids-as potentially pivotal in relaying signals from impaired genomes to the inner mitochondrial membrane. In summary, our study delineates the cascade connecting damaged mitochondrial genomes to the cytoplasm and highlights the significance of the ISR in maintaining mitochondrial homeostasis amid genome instability.

Hyperubiquitylation of DNA helicase RECQL4 by E3 ligase MITOL prevents mitochondrial entry and potentiates mitophagy

Mutations in the DNA helicase RECQL4 lead to Rothmund-Thomson Syndrome (RTS), a disorder characterized by mitochondrial dysfunctions, premature aging, and genomic instability. However, the mechanisms by which these mutations lead to pathology are unclear. Here we report that RECQL4 is ubiquitylated by a mitochondrial E3 ligase, MITOL, at two lysine residues (K1101, K1154) via K6 linkage. This ubiquitylation hampers the interaction of RECQL4 with mitochondrial importer Tom20, thereby restricting its own entry into mitochondria. We show the RECQL4 2K mutant (where both K1101 and K1154 are mutated) has increased entry into mitochondria and demonstrates enhanced mtDNA replication. We observed that the three tested RTS patient mutants were unable to enter the mitochondria and showed decreased mtDNA replication. Furthermore, we found that RECQL4 in RTS patient mutants are hyper-ubiquitylated by MITOL and form insoluble aggregate-like structures on the outer mitochondrial surface. However, depletion of MITOL allows RECQL4 expressed in these RTS mutants to enter mitochondria and rescue mtDNA replication. Finally, we show increased accumulation of hyper-ubiquitylated RECQL4 outside the mitochondria leads to the cells being potentiated to increased mitophagy. Hence, we conclude regulating the turnover of RECQL4 by MITOL may have a therapeutic effect in RTS patients.

Cooperative sensing of mitochondrial DNA by ZBP1 and cGAS promotes cardiotoxicity

Mitochondrial DNA (mtDNA) is a potent agonist of the innate immune system; however, the exact immunostimulatory features of mtDNA and the kinetics of detection by cytosolic nucleic acid sensors remain poorly defined. Here, we show that mitochondrial genome instability promotes Z-form DNA accumulation. Z-DNA binding protein 1 (ZBP1) stabilizes Z-form mtDNA and nucleates a cytosolic complex containing cGAS, RIPK1, and RIPK3 to sustain STAT1 phosphorylation and type I interferon (IFN-I) signaling. Elevated Z-form mtDNA, ZBP1 expression, and IFN-I signaling are observed in cardiomyocytes after exposure to Doxorubicin, a first-line chemotherapeutic agent that induces frequent cardiotoxicity in cancer patients. Strikingly, mice lacking ZBP1 or IFN-I signaling are protected from Doxorubicin-induced cardiotoxicity. Our findings reveal ZBP1 as a cooperative partner for cGAS that sustains IFN-I responses to mitochondrial genome instability and highlight ZBP1 as a potential target in heart failure and other disorders where mtDNA stress contributes to interferon-related pathology.

DNA-protein cross-links between abasic DNA damage and mitochondrial transcription factor A (TFAM).

In higher eukaryotic cells, mitochondria are essential organelles for energy production, metabolism, and signaling. Mitochondrial DNA (mtDNA) encodes 13 protein subunits for oxidative phosphorylation and a set of tRNAs and rRNAs. mtDNA damage, sourced from endogenous chemicals and environmental factors, contributes to mitochondrial genomic instability, which has been associated with various mitochondrial diseases. DNA-protein cross-links (DPCs) are deleterious DNA lesions that threaten genomic integrity. Although much has been learned about the formation and repair of DPCs in the nucleus, little is known about DPCs in mitochondria. Here, we present in vitro and in cellulo data to demonstrate the formation of DPCs between a prevalent abasic (AP) DNA lesion and a DNA-packaging protein, mitochondrial transcription factor A (TFAM). TFAM cleaves AP-DNA and forms DPCs and single-strand breaks (SSB). Lys residues of TFAM are critical for the formation of TFAM-DPC and a reactive 3'-phospho-α,β-unsaturated aldehyde (3'pUA) residue on SSB. The 3'pUA residue reacts with two Cys of TFAM and contributes to the stable TFAM-DPC formation. Glutathione reacts with 3'pUA and competes with TFAM-DPC formation, corroborating our cellular experiments showing the accumulation of TFAM-DPCs under limiting glutathione. Our data point to the involvement of TFAM in AP-DNA turnover and fill a knowledge gap regarding the protein factors in processing damaged mtDNA.

Unleashing a novel function of Endonuclease G in mitochondrial genome instability.

Having its genome makes the mitochondrion a unique and semiautonomous organelle within cells. Mammalian mitochondrial DNA (mtDNA) is a double-stranded closed circular molecule of about 16 kb coding for 37 genes. Mutations, including deletions in the mitochondrial genome, can culminate in different human diseases. Mapping the deletion junctions suggests that the breakpoints are generally seen at hotspots. '9 bp deletion' (8271-8281), seen in the intergenic region of cytochrome c oxidase II/tRNALys, is the most common mitochondrial deletion. While it is associated with several diseases like myopathy, dystonia, and hepatocellular carcinoma, it has also been used as an evolutionary marker. However, the mechanism responsible for its fragility is unclear. In the current study, we show that Endonuclease G, a mitochondrial nuclease responsible for nonspecific cleavage of nuclear DNA during apoptosis, can induce breaks at sequences associated with '9 bp deletion' when it is present on a plasmid or in the mitochondrial genome. Through a series of in vitro and intracellular studies, we show that Endonuclease G binds to G-quadruplex structures formed at the hotspot and induces DNA breaks. Therefore, we uncover a new role for Endonuclease G in generating mtDNA deletions, which depends on the formation of G4 DNA within the mitochondrial genome. In summary, we identify a novel property of Endonuclease G, besides its role in apoptosis and the recently described 'elimination of paternal mitochondria during fertilisation.

Sarcopenia: An Age-Related Multifactorial Disorder.

Sarcopenia is a disease characterized by a gradual decline in skeletal muscle mass and strength that accompanies the normal aging process. The disease is associated with various adverse health outcomes in the geriatric population, such as prolonged hospital admission, disability, poor quality of life, frailty, and mortality. Factors involved in the development of age-related sarcopenia include anorexia, hormone level alterations, decreased neural innervation, low muscular blood flow, cytokine dysregulation, altered mitochondrial activity, genomic instability, intracellular proteolysis, and insulin resistance. Understanding the underlying mechanisms may facilitate the development of efficient prophylactic and therapeutic strategies to improve the quality of life in elderly individuals. Thus, the present review highlights literature regarding the mechanism of sarcopenia development in elderly patients.

Mitochondrial DNA Mutagenesis: Feature of and Biomarker for Environmental Exposures and Aging.

Mitochondrial dysfunction is a hallmark of aging. Mitochondrial genome (mtDNA) instability contributes to mitochondrial dysfunction, and mtDNA mutagenesis may contribute to aging. However, the origin of mtDNA mutations remains somewhat controversial. The goals of this review are to introduce and review recent literature on mtDNA mutagenesis and aging, address recent animal and epidemiological evidence for the effects of chemicals on mtDNA damage and mutagenesis, propose hypotheses regarding the contribution of environmental toxicant exposure to mtDNA mutagenesis in the context of aging, and suggest future directions and approaches for environmental health researchers. Stressors such as pollutants, pharmaceuticals, and ultraviolet radiation can damage the mitochondrial genome or disrupt mtDNA replication, repair, and organelle homeostatic processes, potentially influencing the rate of accumulation of mtDNA mutations. Accelerated mtDNA mutagenesis could contribute to aging, diseases of aging, and sensitize individuals with pathogenic mtDNA variants to stressors. We propose three potential mechanisms of toxicant-induced effects on mtDNA mutagenesis over lifespan: (1) increased de novo mtDNA mutations, (2) altered frequencies of mtDNA mutations, or (3) both. There are remarkably few studies that have investigated the impact of environmental chemical exposures on mtDNA instability and mutagenesis, and even fewer in the context of aging. More studies are warranted because people are exposed to tens of thousands of chemicals, and are living longer. Finally, we suggest that toxicant-induced mtDNA damage and mutational signatures may be a sensitive biomarker for some exposures.

Base excision repair causes age-dependent accumulation of single-stranded DNA breaks that contribute to Parkinson disease pathology.

SummaryAging, genomic stress, and mitochondrial dysfunction are risk factors for neurodegenerative pathologies, such as Parkinson disease (PD). Although genomic instability is associated with aging and mitochondrial impairment, the underlying mechanisms are poorly understood. Here, we show that base excision repair generates genomic stress, promoting age-related neurodegeneration in a Caenorhabditis elegans PD model. A physiological level of NTH-1 DNA glycosylase mediates mitochondrial and nuclear genomic instability, which promote degeneration of dopaminergic neurons in older nematodes. Conversely, NTH-1 deficiency protects against α-synuclein-induced neurotoxicity, maintaining neuronal function with age. This apparent paradox is caused by modulation of mitochondrial transcription in NTH-1-deficient cells, and this modulation activates LMD-3, JNK-1, and SKN-1 and induces mitohormesis. The dependance of neuroprotection on mitochondrial transcription highlights the integration of BER and transcription regulation during physiological aging. Finally, whole-exome sequencing of genomic DNA from patients with idiopathic PD suggests that base excision repair might modulate susceptibility to PD in humans.

G-quadruplexes in the mitochondrial genome - a cause for instability.

Accumulation of mutations such as deletions in mitochondrial DNA is associated with ageing, cancer and human genetic disorders. These deletions are often flanked by GC-skewed sequence motifs that can potentially fold into secondary non-B DNA conformations. G-quadruplexes are emerging as key initiators of mitochondrial genomic instability. In this issue, Dahal etal provide an insilico analysis of sequence motifs that can fold into altered DNA structures in mitochondrial genomic regions that contain frequent deletions. They show the formation of five G-quadruplexes near such frequent breakpoints using biochemical and biophysical approaches invitro and more importantly inside mammalian cells. Comment on: https://doi.org/10.1111/febs.16113.

How Inflammation Pathways Contribute to Cell Death in Neuro-Muscular Disorders.

Neuro-muscular disorders include a variety of diseases induced by genetic mutations resulting in muscle weakness and waste, swallowing and breathing difficulties. However, muscle alterations and nerve depletions involve specific molecular and cellular mechanisms which lead to the loss of motor-nerve or skeletal-muscle function, often due to an excessive cell death. Morphological and molecular studies demonstrated that a high number of these disorders seem characterized by an upregulated apoptosis which significantly contributes to the pathology. Cell death involvement is the consequence of some cellular processes that occur during diseases, including mitochondrial dysfunction, protein aggregation, free radical generation, excitotoxicity and inflammation. The latter represents an important mediator of disease progression, which, in the central nervous system, is known as neuroinflammation, characterized by reactive microglia and astroglia, as well the infiltration of peripheral monocytes and lymphocytes. Some of the mechanisms underlying inflammation have been linked to reactive oxygen species accumulation, which trigger mitochondrial genomic and respiratory chain instability, autophagy impairment and finally neuron or muscle cell death. This review discusses the main inflammatory pathways contributing to cell death in neuro-muscular disorders by highlighting the main mechanisms, the knowledge of which appears essential in developing therapeutic strategies to prevent the consequent neuron loss and muscle wasting.

Characterization of G4 DNA formation in mitochondrial DNA and their potential role in mitochondrial genome instability.

Mitochondria possess their own genome which can be replicated independently of nuclear DNA. Mitochondria being the powerhouse of the cell produce reactive oxygen species, due to which the mitochondrial genome is frequently exposed to oxidative damage. Previous studies have demonstrated an association of mitochondrial deletions to aging and human disorders. Many of these deletions were present adjacent to non-B DNA structures. Thus, we investigate noncanonical structures associated with instability in mitochondrial genome. Insilico studies revealed the presence of >100 G-quadruplex motifs (of which 5 have the potential to form 3-plate G4 DNA), 23 inverted repeats, and 3 mirror repeats in the mitochondrial DNA (mtDNA). Further analysis revealed that among the deletion breakpoints from patients with mitochondrial disorders, majority are located at G4 DNA motifs. Interestingly, ~50% of the deletions were at base-pair positions 8271-8281, ~35% were due to deletion at 12362-12384, and ~12% due to deletion at 15516-15545. Formation of 3-plate G-quadruplex DNA structures at mitochondrial fragile regions was characterized using electromobility shift assay, circular dichroism (CD), and Taq polymerase stop assay. All 5 regions could fold into both intramolecular and intermolecular G-quadruplex structures in a KCl-dependent manner. G4 DNA formation was in parallel orientation, which was abolished in the presence of LiCl. The formation of G4 DNA affected both replication and transcription. Finally, immunolocalization of BG4 with MitoTracker confirmed the formation of G-quadruplex in mitochondrial genome. Thus, we characterize the formation of 5 different G-quadruplex structures in human mitochondrial region, which may contribute toward formation of mitochondrial deletions.

Nuclear and mitochondrial genome instability induced by fractions of ethanolic extract from Hovenia dulcis Thunberg in Saccharomyces cerevisiae strains.

Hovenia dulcis is a plant commonly used as a pharmaceutical supplement, having displayed important pharmacological properties such antigiardic, antineoplastic and hepatoprotective. The purpose of this work was investigate the cytotoxic, genotoxic and mutagenic potential from fractions of Hovenia dulcis ethanolic extract on Saccharomyces cerevisiae strains FF18733 (wild type) and CD138 (ogg1). Ethanolic extract from Hovenia dulcis leaves was fractioned using organic solvents according to increasing polarity: Hexane (1:1), dichlorometane (1:1), ethyl acetate (1:1) and butanol (1:1). Three experimental assays were performed, such as (i) inactivation of cultures; (ii) mutagenesis (canavanine resistance system) and (iii) loss of mitochondrial function (petites colonies). The findings shown a decrease in cell viability in FF18733 and CD138 strains; all fractions of the extract were mutagenic in CD138 strain; only ethyl acetate and butanol fractions increased the rate of petites colonies for CD138 strains. Ethyl acetate and n-butanol fractions induces mutagenicity, at the evaluated concentrations, in mitochondrial and genomic DNA in CD138 strain, mediated by oxidative lesions. In conclusion, it is possible to infer that the lesions caused by the extract fractions could be mediated by reactive oxygen species and might reach multiple molecular targets to cause cellular damage.

Muscle Mitochondrial DNA Copy Number, Deletion Mutation Frequency, and Physical Performance in Older Adults

Mitochondrial DNA (mtDNA) quantity and quality influence hallmarks of aging – mitochondrial dysfunction and genomic instability. The interactions between mtDNA quantity and quality and physical performance have not been extensively examined in humans. The aim of this study was to test the interactions between skeletal muscle mtDNA copy number, mtDNA deletion mutation frequency, and physical performance measures in older adults. Total DNA was isolated from muscle biopsies and used for quantitation of mtDNA copy number and mutation frequency by digital PCR. The biopsies were obtained from a cross-sectional cohort of 53 adults aged 50 to 86 years. Before the biopsy, physical performance measures were collected. MtDNA deletions increased exponentially with advancing age. On average, mtDNA deletion frequency increased 18-fold between 50 and 80, with a trend toward lower deletion frequency in females. MtDNA deletion frequency predicted declines in VO2 max, where 4.7% of the variation in VO2 max was explained by mtDNA deletion frequency. MtDNA copy number was negatively correlated with age and mtDNA deletion frequency, but positively correlated with lean mass. There was a trend to lower mtDNA deletion frequency in females, consistent with increased longevity in females. Larger studies may better delineate sex effects. These data are consistent with a role for mitochondrial function and genome integrity in the maintenance of physical performance with age. Analyses of mtDNA quality and quantity in longitudinal studies could extend our understanding of the importance of mitochondria in human aging and longevity.

The Impact of Mitochondrial Deficiencies in Neuromuscular Diseases.

Neuromuscular diseases (NMDs) are a heterogeneous group of acquired or inherited rare disorders caused by injury or dysfunction of the anterior horn cells of the spinal cord (lower motor neurons), peripheral nerves, neuromuscular junctions, or skeletal muscles leading to muscle weakness and waste. Unfortunately, most of them entail serious or even fatal consequences. The prevalence rates among NMDs range between 1 and 10 per 100,000 population, but their rarity and diversity pose difficulties for healthcare and research. Some molecular hallmarks are being explored to elucidate the mechanisms triggering disease, to set the path for further advances. In fact, in the present review we outline the metabolic alterations of NMDs, mainly focusing on the role of mitochondria. The aim of the review is to discuss the mechanisms underlying energy production, oxidative stress generation, cell signaling, autophagy, and inflammation triggered or conditioned by the mitochondria. Briefly, increased levels of inflammation have been linked to reactive oxygen species (ROS) accumulation, which is key in mitochondrial genomic instability and mitochondrial respiratory chain (MRC) dysfunction. ROS burst, impaired autophagy, and increased inflammation are observed in many NMDs. Increasing knowledge of the etiology of NMDs will help to develop better diagnosis and treatments, eventually reducing the health and economic burden of NMDs for patients and healthcare systems.

THU045 - Non-alcoholic steatohepatitis is associated with liver mitochondrial DNA damage and genome instability of cytochrome b, a component of the cellular respirasome supercomplex

THU045 - Non-alcoholic steatohepatitis is associated with liver mitochondrial DNA damage and genome instability of cytochrome b, a component of the cellular respirasome supercomplex

NME3 Regulates Mitochondria to Reduce ROS-Mediated Genome Instability.

NME3 is a member of the nucleoside diphosphate kinase (NDPK) family that binds to the mitochondrial outer membrane to stimulate mitochondrial fusion. In this study, we showed that NME3 knockdown delayed DNA repair without reducing the cellular levels of nucleotide triphosphates. Further analyses revealed that NME3 knockdown increased fragmentation of mitochondria, which in turn led to mitochondrial oxidative stress-mediated DNA single-strand breaks (SSBs) in nuclear DNA. Re-expression of wild-type NME3 or inhibition of mitochondrial fission markedly reduced SSBs and facilitated DNA repair in NME3 knockdown cells, while expression of N-terminal deleted mutant defective in mitochondrial binding had no rescue effect. We further showed that disruption of mitochondrial fusion by knockdown of NME4 or MFN1 also caused mitochondrial oxidative stress-mediated genome instability. In conclusion, the contribution of NME3 to redox-regulated genome stability lies in its function in mitochondrial fusion.

Abstract 4907: Transcriptomic profiling identifies a novel mitochondrial gene expression signature that predicts recurrence in microsatellite stable stage II and III colorectal cancer

Abstract Purpose: Colorectal cancer (CRC) is the third most commonly diagnosed cancer in the United States. Current TNM (Tumor, Node, and Metastasis) classification approach is suboptimal in determining the prognosis of CRC patients. While there is a general consensus that stage II and III CRC patients with microsatellite instability (MSI) exhibit better outcomes, there is a lack of availability of biomarkers that can predict prognosis in microsatellite stable (MSS) patients. Given the emerging evidence that mitochondrial (mt) genomic instability is one of the hallmark features of cancer progression, herein, we systematically explored and established a novel mt-gene panel as a prognostic signature in CRC. Experimental design: Three genome wide mRNA expression profiling datasets were analyzed for the biomarker discovery (GSE33113, n=65) and validation (GSE14333, n=196; TCGA, n=62) of a mt-gene expression signature that predicts recurrence in stage II/III MSS CRC patients. Subsequently, a 14-gene signature was established, and independently tested (n=257) and validated (n=212) in two clinical cohorts, using qRT-PCR assays. The uni- and multi-variate Cox proportional hazard (CoxPH) regression analyses were performed to evaluate the performance of the mt-gene signature individually, and in combination with various clinicopathological factors (e.g. TNM staging, lymphatic invasion, and tumor infiltration) for predicting recurrence free survival (RFS). Results: The expression analysis of 568 mt-genes in CRC patients with and without recurrence, resulted in the identification of 32 significantly dysregulated genes (p<0.05). The univariate CoxPH analysis yielded a 14-gene signature that accurately predicted recurrence in the discovery (AUC = 0.89), and two independent validation cohorts (AUC = 0.75 and 0.79, respectively). This 14-gene signature demonstrated significant associations with poor RFS in independent testing and clinical validation cohorts of CRC patients (AUC = 0.87 and 0.73, respectively). Univariate analysis revealed that in addition to the mt-gene signature, TNM-stage (p = 0.007) and lymphatic invasion (p = 0.0004) were significant predictors of RFS; while multivariate analyses demonstrated that our mt-gene signature was an independent predictor (HR = 2.71; P<0.001) of tumor recurrence in stage II/III CRC patients. Intriguingly, a risk-stratification model comprising of the 14-gene signature together with TNM-stage and lymphatic invasion status achieved an even superior AUC of 0.89 and 0.78 in both clinical cohorts. Conclusions: We have identified a novel, 14-gene signature as an independent predictor of tumor recurrence; which in combination with TNM-stage and lymphatic invasion, offers an easy-to-translate and facile assay for the personalized risk-assessment in stage II/III MSS-CRC patients. Citation Format: SOUVIK GHATAK, Raju Kandimalla, Jasjit K. Banwait, Hiroyuki Uetake, Francesc Balaguer, Luis Bujanda, Ajay Goel. Transcriptomic profiling identifies a novel mitochondrial gene expression signature that predicts recurrence in microsatellite stable stage II and III colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4907.

Prevention of mitochondrial genomic instability in yeast by the mitochondrial recombinase Mhr1

Mitochondrial (mt) DNA encodes factors essential for cellular respiration, therefore its level and integrity are crucial. ABF2 encodes a mitochondrial DNA-binding protein and its null mutation (Δabf2) induces mtDNA instability in Saccharomyces cerevisiae. Mhr1 is a mitochondrial recombinase that mediates the predominant form of mtDNA replication and acts in mtDNA segregation and the repair of mtDNA double-stranded breaks (DSBs). However, the involvement of Mhr1 in prevention of mtDNA deletion mutagenesis is unknown. In this study we used Δabf2 mhr1-1 double-mutant cells, which lose mitochondrial function in media containing fermentable carbon sources, to investigate whether Mhr1 is a suppressor of mtDNA deletion mutagenesis. We used a suppresivity assay and Southern blot analysis to reveal that the Δabf2 mutation causes mtDNA deletions rather than an mtDNA-lacking (ρ0) phenotype, and observed that mtDNA deletions are exacerbated by an additional mhr1-1 mutation. Loss of respiratory function due to mtDNA fragmentation occurred in ∆mhr1 and ∆abf2 mhr1-1 cells. However, exogenous introduction of Mhr1 into Δabf2 mhr1-1 cells significantly rescued respiratory growth, suggesting that Mhr1-driven homologous mtDNA recombination prevents mtDNA instability.

Mitochondrial DNA Fork Protection by Fanconi Anemia Genes Suppresses cGAS Activation

Fanconi Anemia patients exhibit bone marrow failure, increased cancer risk, developmental abnormalities, endocrine dysfunction and elevated inflammation. The underlying cause for these pleiotropic disease phenotypes remains enigmatic. Here, we show that Fanconi Anemia genes are required for protecting mitochondrial DNA replication forks upon oxidative damage to suppress mitochondrial genome instability. Specifically, patient cells show a loss of nascent mtDNA caused by MRE11 nuclease degradation. Mitochondrial fork protection requires multiple pathway membersincluding FANCD2, RAD51C, BRCA2, and SLX4, but not Fanconi Anemia core complex genes or FANCD2 mono-ubiquitination, revealing a mechanistic separation between mitochondrial and nuclear FANC genome stability pathways. Importantly, stalled mitochondrial replication fork degradation by MRE11 results in activation of cGAS, thus identifying a nuclease responsible for mitochondrial DNA-dependent induction of the cGAS-inflammation pathway. The collective results establish a molecular pathway for mitochondrial DNA replication stability and explain how defects in genome maintenance proteins cause diverse cellular disease phenotypes.

Mitochondrial genome analysis in penile carcinoma.

Penile cancer is a rare neoplasm that seems to be linked to socio-economic differences. Mitochondrial genome alterations are common in many tumors types and are reported as regulating oxidative metabolism and impacting tumorigenesis. In this study, we evaluate for the first time the mitochondrial genome in penile carcinoma (PeCa), aiming to evaluate heteroplasmy, mitochondrial DNA (mtDNA) mutational load and mtDNA content in Penile tumors. Using next generation sequencing (NGS), we sequenced the mitochondrial genome of 13 penile tumors and 12 non-neoplastic tissue samples, which allowed us to identify mtDNA variants and heteroplasmy. We further evaluated variant's pathogenicity using Mutpred predictive software and calculated mtDNA content using quantitative PCR. Mitochondrial genome sequencing revealed an increase number of non-synonymous variants in the tumor tissue, along with higher frequency of heteroplasmy and mtDNA depletion in penile tumors, suggesting an increased mitochondrial instability in penile tumors. We also described a list of mitochondrial variants found in penile tumor and normal tissue, including five novel variants found in the tumoral tissue. Our results showed an increased mitochondrial genome instability in penile tumors. We also suggest that mitochondrial DNA copy number (mtDNAcn) and mtDNA variants may act together to imbalance mitochondrial function in PeCa. The better understanding of mitochondrial biology can bring new insights on mechanisms and open a new field for therapy in PeCa.