RESUMO
All metazoans depend on the consumption of O2 by the mitochondrial oxidative phosphorylation system (OXPHOS) to produce energy. In addition, the OXPHOS uses O2 to produce reactive oxygen species that can drive cell adaptations1-4, a phenomenon that occurs in hypoxia4-8 and whose precise mechanism remains unknown. Ca2+ is the best known ion that acts as a second messenger9, yet the role ascribed to Na+ is to serve as a mere mediator of membrane potential10. Here we show that Na+ acts as a second messenger that regulates OXPHOS function and the production of reactive oxygen species by modulating the fluidity of the inner mitochondrial membrane. A conformational shift in mitochondrial complex I during acute hypoxia11 drives acidification of the matrix and the release of free Ca2+ from calcium phosphate (CaP) precipitates. The concomitant activation of the mitochondrial Na+/Ca2+ exchanger promotes the import of Na+ into the matrix. Na+ interacts with phospholipids, reducing inner mitochondrial membrane fluidity and the mobility of free ubiquinone between complex II and complex III, but not inside supercomplexes. As a consequence, superoxide is produced at complex III. The inhibition of Na+ import through the Na+/Ca2+ exchanger is sufficient to block this pathway, preventing adaptation to hypoxia. These results reveal that Na+ controls OXPHOS function and redox signalling through an unexpected interaction with phospholipids, with profound consequences for cellular metabolism.
Assuntos
Transporte de Elétrons , Hipóxia/metabolismo , Mitocôndrias/metabolismo , Sistemas do Segundo Mensageiro , Sódio/metabolismo , Animais , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Fosfatos de Cálcio/metabolismo , Linhagem Celular Tumoral , Precipitação Química , Humanos , Masculino , Fluidez de Membrana , Camundongos Endogâmicos C57BL , Membranas Mitocondriais/química , Membranas Mitocondriais/metabolismo , Proteínas Mitocondriais/metabolismo , Fosforilação Oxidativa , Ratos , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismo , Trocador de Sódio e Cálcio/metabolismoRESUMO
BACKGROUND: In most eukaryotic cells, the mitochondrial DNA (mtDNA) is transmitted uniparentally and present in multiple copies derived from the clonal expansion of maternally inherited mtDNA. All copies are therefore near-identical, or homoplasmic. The presence of >1 mtDNA variant in the same cytoplasm can arise naturally or result from new medical technologies aimed at preventing mitochondrial genetic diseases and improving fertility. The latter is called divergent nonpathologic mtDNA heteroplasmy (DNPH). We hypothesized that DNPH is maladaptive and usually prevented by the cell. METHODS: We engineered and characterized DNPH mice throughout their lifespan using transcriptomic, metabolomic, biochemical, physiologic, and phenotyping techniques. We focused on in vivo imaging techniques for noninvasive assessment of cardiac and pulmonary energy metabolism. RESULTS: We show that DNPH impairs mitochondrial function, with profound consequences in critical tissues that cannot resolve heteroplasmy, particularly cardiac and skeletal muscle. Progressive metabolic stress in these tissues leads to severe pathology in adulthood, including pulmonary hypertension and heart failure, skeletal muscle wasting, frailty, and premature death. Symptom severity is strongly modulated by the nuclear context. CONCLUSIONS: Medical interventions that may generate DNPH should address potential incompatibilities between donor and recipient mtDNA.
Assuntos
Fragilidade , Cardiopatias , Hipertensão Pulmonar , Adulto , Animais , DNA Mitocondrial/genética , Fragilidade/patologia , Cardiopatias/patologia , Heteroplasmia , Humanos , Hipertensão Pulmonar/genética , Hipertensão Pulmonar/patologia , Camundongos , Mitocôndrias/genéticaRESUMO
Human mitochondrial DNA (mtDNA) shows extensive within population sequence variability. Many studies suggest that mtDNA variants may be associated with ageing or diseases, although mechanistic evidence at the molecular level is lacking. Mitochondrial replacement has the potential to prevent transmission of disease-causing oocyte mtDNA. However, extension of this technology requires a comprehensive understanding of the physiological relevance of mtDNA sequence variability and its match with the nuclear-encoded mitochondrial genes. Studies in conplastic animals allow comparison of individuals with the same nuclear genome but different mtDNA variants, and have provided both supporting and refuting evidence that mtDNA variation influences organismal physiology. However, most of these studies did not confirm the conplastic status, focused on younger animals, and did not investigate the full range of physiological and phenotypic variability likely to be influenced by mitochondria. Here we systematically characterized conplastic mice throughout their lifespan using transcriptomic, proteomic,metabolomic, biochemical, physiological and phenotyping studies. We show that mtDNA haplotype profoundly influences mitochondrial proteostasis and reactive oxygen species generation,insulin signalling, obesity, and ageing parameters including telomere shortening and mitochondrial dysfunction, resulting in profound differences in health longevity between conplastic strains.
Assuntos
Envelhecimento/genética , Núcleo Celular/genética , DNA Mitocondrial/genética , Variação Genética/genética , Metabolismo/genética , Mitocôndrias/genética , Mitocôndrias/metabolismo , Envelhecimento/fisiologia , Animais , Feminino , Genoma Mitocondrial/genética , Haplótipos , Insulina/metabolismo , Longevidade/genética , Masculino , Metabolismo/fisiologia , Metabolômica , Camundongos , Camundongos Congênicos , Mitocôndrias/patologia , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Obesidade/genética , Obesidade/metabolismo , Fenótipo , Proteômica , Espécies Reativas de Oxigênio/metabolismo , Encurtamento do Telômero , Transcriptoma , Resposta a Proteínas não DobradasRESUMO
In the past decade, anti-tumour immune responses have been successfully exploited to improve the outcome of patients with different cancers. Significant progress has been made in taking advantage of different types of T cell functions for therapeutic purposes. Despite these achievements, only a subset of patients respond favorably to immunotherapy. Therefore, there is a need of novel approaches to improve the effector functions of immune cells and to recognize the major targets of anti-tumour immunity. A major hallmark of cancer is metabolic rewiring associated with switch of mitochondrial functions. These changes are a consequence of high energy demand and increased macromolecular synthesis in cancer cells. Such adaptations in tumour cells might generate novel targets of tumour therapy, including the generation of neoantigens. Here, we review the most recent advances in research on the immune response to mitochondrial proteins in different cellular conditions.
Assuntos
Proteínas Mitocondriais , Neoplasias , Antígenos de Neoplasias , Humanos , Imunoterapia , Neoplasias/terapia , Linfócitos TRESUMO
The oxidative phosphorylation (OXPHOS) system is the only structure in animal cells with components encoded by two genomes, maternally transmitted mitochondrial DNA (mtDNA), and biparentally transmitted nuclear DNA (nDNA). MtDNA-encoded genes have to physically assemble with their counterparts encoded in the nucleus to build together the functional respiratory complexes. Therefore, structural and functional matching requirements between the protein subunits of these molecular complexes are rigorous. The crosstalk between nDNA and mtDNA needs to overcome some challenges, as the nuclear-encoded factors have to be imported into the mitochondria in a correct quantity and match the high number of organelles and genomes per mitochondria that encode and synthesize their own components locally. The cell is able to sense the mito-nuclear match through changes in the activity of the OXPHOS system, modulation of the mitochondrial biogenesis, or reactive oxygen species production. This implies that a complex signaling cascade should optimize OXPHOS performance to the cellular-specific requirements, which will depend on cell type, environmental conditions, and life stage. Therefore, the mitochondria would function as a cellular metabolic information hub integrating critical information that would feedback the nucleus for it to respond accordingly. Here, we review the current understanding of the complex interaction between mtDNA and nDNA.
Assuntos
Núcleo Celular/genética , DNA Mitocondrial , Fosforilação Oxidativa , Animais , Citoplasma/genética , Citoplasma/metabolismo , DNA Mitocondrial/genética , DNA Mitocondrial/metabolismo , Evolução Molecular , Regulação da Expressão Gênica , Genoma , Haplótipos , Humanos , Mitocôndrias/genéticaRESUMO
Cardiac progenitors are specified early in development and progressively differentiate and mature into fully functional cardiomyocytes. This process is controlled by an extensively studied transcriptional program. However, the regulatory events coordinating the progression of such program from development to maturation are largely unknown. Here, we show that the genome organizer CTCF is essential for cardiogenesis and that it mediates genomic interactions to coordinate cardiomyocyte differentiation and maturation in the developing heart. Inactivation of Ctcf in cardiac progenitor cells and their derivatives in vivo during development caused severe cardiac defects and death at embryonic day 12.5. Genome wide expression analysis in Ctcf mutant hearts revealed that genes controlling mitochondrial function and protein production, required for cardiomyocyte maturation, were upregulated. However, mitochondria from mutant cardiomyocytes do not mature properly. In contrast, multiple development regulatory genes near predicted heart enhancers, including genes in the IrxA cluster, were downregulated in Ctcf mutants, suggesting that CTCF promotes cardiomyocyte differentiation by facilitating enhancer-promoter interactions. Accordingly, loss of CTCF disrupts gene expression and chromatin interactions as shown by chromatin conformation capture followed by deep sequencing. Furthermore, CRISPR-mediated deletion of an intergenic CTCF site within the IrxA cluster alters gene expression in the developing heart. Thus, CTCF mediates local regulatory interactions to coordinate transcriptional programs controlling transitions in morphology and function during heart development.
Assuntos
Cromatina/genética , Desenvolvimento Embrionário/genética , Ventrículos do Coração/crescimento & desenvolvimento , Coração/crescimento & desenvolvimento , Proteínas Repressoras/genética , Animais , Fator de Ligação a CCCTC , Diferenciação Celular/genética , Elementos Facilitadores Genéticos , Regulação da Expressão Gênica no Desenvolvimento , Coração/embriologia , Ventrículos do Coração/embriologia , Camundongos , Mitocôndrias/genética , Mitocôndrias/metabolismo , Organogênese/genética , Regiões Promotoras Genéticas , Ligação Proteica , Ativação Transcricional/genéticaRESUMO
We report the synthesis of micellar phosphatidylcholine-coated superparamagnetic iron oxide nanoparticles as a new long circulation contrast agents for magnetic resonance imaging. Oleic acid-coated Fe3 O4 nanoparticles were first prepared through thermal degradation and then encapsulated into small clusters with a phosphatidylcholine coating to obtain hydrophilic nanomicelles. A thorough characterization confirmed the chemical nature of the coating and the excellent colloidal stability of these nanomicelles in aqueous media. Magnetization and relaxivity properties proved their suitability as magnetic resonance imaging (MRI) contrast agent and in vitro cell viability data showed low toxicity. Vascular lifetime and elimination kinetics in the liver were assessed by blood relaxometry and by in vivo MRI in rats and compared with "control" particles prepared with a polyethylene glycol derivative. These micellar particles had a lifetime in blood of more than 10â h, much longer than the control nanoparticles (≈2â h), which is remarkable considering that the coating molecule is a small biocompatible zwitterionic phospholipid. The protein corona was characterized after incubation with rat serum at different times by high-throughput proteomics, showing a higher proportion of bound apolipoproteins and other dysopsonins for the phosphatidylcholine particles. The antibiofouling properties of this corona and its resistance to the adsorption of proteins corroborate the observed enhanced stability and prolonged systemic circulation.
Assuntos
Meios de Contraste/química , Compostos Férricos/química , Nanopartículas de Magnetita/química , Fosfatidilcolinas/química , Animais , Linhagem Celular , Meios de Contraste/farmacocinética , Compostos Férricos/sangue , Imageamento por Ressonância Magnética , Masculino , Camundongos Endogâmicos C57BL , Micelas , Fosfatidilcolinas/sangue , Ratos , Ratos WistarRESUMO
IFNγ is an immune mediator with concomitant pro- and anti-tumor functions. Here, we provide evidence that IFNγ directly acts on intra-tumoral CD8 T cells to restrict anti-tumor responses. We report that expression of the IFNγ receptor ß chain (IFNγR2) in CD8 T cells negatively correlates with clinical responsiveness to checkpoint blockade in metastatic melanoma patients, suggesting that the loss of sensitivity to IFNγ contributes to successful antitumor immunity. Indeed, specific deletion of IFNγR in CD8 T cells promotes tumor control in a mouse model of melanoma. Chronic IFNγ inhibits the maintenance, clonal diversity and proliferation of stem-like T cells. This leads to decreased generation of T cells with intermediate expression of exhaustion markers, previously associated with beneficial anti-tumor responses. This study provides evidence of a negative feedback loop whereby IFNγ depletes stem-like T cells to restrict anti-tumor immunity. Targeting this pathway might represent an alternative strategy to enhance T cell-based therapies.
Assuntos
Melanoma , Linfócitos T Citotóxicos , Camundongos , Animais , Linfócitos T Citotóxicos/metabolismo , Linfócitos T CD8-Positivos , Melanoma/terapia , Melanoma/tratamento farmacológico , Células Clonais/metabolismoRESUMO
Cancer heterogeneity and evolution are not fully understood. Here, we show that mitochondrial DNA of the normal liver shapes tumor progression, histology, and immune environment prior to the acquisition of oncogenic mutation. Using conplastic mice, we show that mtDNA dictates the expression of the mitochondrial unfolded protein response (UPRmt) in the normal liver. Activation of oncogenic mutations in UPRmt-positive liver increases tumor incidence and histological heterogeneity. Further, in a subset of UPRmt-positive mice, invasive liver cancers develop. RNA sequencing (RNA-seq) analysis of the normal liver reveals that, in this subset, the PAPP-A/DDR2/SNAIL axis of invasion pre-exists along with elevated collagen. Since PAPP-A promotes immune evasion, we analyzed the immune signature and found that their livers are immunosuppressed. Further, the PAPP-A signature identifies the immune exhausted subset of hepatocellular carcinoma (HCC) in humans. Our data suggest that mtDNA of normal liver shapes the entire liver cancer portrait upon acquisition of oncogenic mutations.
Assuntos
Carcinoma Hepatocelular/genética , DNA Mitocondrial/genética , Neoplasias Hepáticas/genética , Resposta a Proteínas não Dobradas/genética , Animais , Carcinoma Hepatocelular/patologia , Feminino , Humanos , Neoplasias Hepáticas/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteína Plasmática A Associada à Gravidez/metabolismo , TranscriptomaRESUMO
Mitochondria and mtDNA variations contribute to specific aspects of the aging process. Here, we aimed to investigate the influence of mtDNA variation on joint damage in a model of aging using conplastic mice. A conplastic (BL/6NZB) mouse strain was developed with the C57BL/6JOlaHsd nuclear genome and NZB/OlaHsd mtDNA, for comparison with the original C57BL/6JOlaHsd strain (BL/6C57). Conplastic (BL/6NZB) and BL/6C57 mice were sacrificed at 25, 75, and 90 weeks of age. Hind knee joints were processed for histological analysis and joint pathology graded using the Mankin scoring system. By immunohistochemistry, cartilage expression of markers of autophagy (LC3, Beclin-1, and P62) and markers of senescence (MMP13, beta-Galactosidase, and p16) and proliferation (Ki67) were analyzed. We also measured the expression of 8-oxo-dG and cleaved caspase-3. Conplastic (BL/6NZB) mice presented lower Mankin scores at 25, 75, and 90 weeks of age, higher expression of LC3 and Beclin-1 and lower of P62 in cartilage than the original strain. Moreover, the downregulation of MMP13, beta-Galactosidase, and p16 was detected in cartilage from conplastic (BL/6NZB) mice, whereas higher Ki67 levels were detected in these mice. Finally, control BL/6C57 mice showed higher cartilage expression of 8-oxo-dG and cleaved caspase-3 than conplastic (BL/6NZB) mice. This study demonstrates that mtDNA genetic manipulation ameliorates joint aging damage in a conplastic mouse model, suggesting that mtDNA variability is a prognostic factor for aging-related osteoarthritis (OA) and that modulation of mitochondrial oxidative phosphorylation (OXPHOS) could be a novel therapeutic target for treating OA associated with aging.
Assuntos
DNA Mitocondrial , Osteoartrite , 8-Hidroxi-2'-Desoxiguanosina , Envelhecimento/fisiologia , Animais , Proteína Beclina-1/genética , Proteína Beclina-1/metabolismo , Caspase 3/metabolismo , DNA Mitocondrial/genética , DNA Mitocondrial/metabolismo , Modelos Animais de Doenças , Antígeno Ki-67/metabolismo , Metaloproteinase 13 da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/metabolismo , Osteoartrite/genética , Osteoartrite/metabolismo , beta-Galactosidase/metabolismoRESUMO
The coating composition of nanomedicines is one of the main features in determining the medicines' fate, clearance, and immunoresponse in the body. To highlight the coatings' impact in pulmonary administration, two micellar superparamagnetic iron oxide nanoparticles (SPION) were compared. These nanoparticles are similar in size and charge but have different coatings: either phosphatidylcholine (PC-SPION) or bovine serum albumin (BSA-SPION). The aim of the study was to increase the understanding of the nano-bio interaction with the cellular and non-cellular components of the lung and underline valuable coatings either for local lung-targeted drug delivery in theranostic application or patient-friendly route systemic administration. PC-SPION and BSA-SPION were deposited in the alveoli by in vivo instillation and, despite the complexity of imaging the lung, SPION were macroscopically visualized by MRI. Impressively, PC-SPION were retained within the lungs for at least a week, while BSA-SPION were cleared more rapidly. The different lung residence times were confirmed by histological analysis and supported by a flow cytometry analysis of the SPION interactions with different myeloid cell populations. To further comprehend the way in which these nanoformulations interact with lung components at the molecular level, we used fluorescence spectroscopy, turbidity measurements, and dynamic light scattering to evaluate the interactions of the two SPION with surfactant protein A (SP-A), a key protein in setting up the nanoparticle behavior in the alveolar fluid. We found that SP-A induced aggregation of PC-SPION, but not BSA-SPION, which likely caused PC-SPION retention in the lung without inducing inflammation. In conclusion, the two SPION show different outcomes from interaction with SP-A leading to distinctive fate in the lung. PC-SPION hold great promise as imaging and theranostic agents when prolonged pulmonary drug delivery is required.
Assuntos
Nanopartículas , Proteína A Associada a Surfactante Pulmonar , Humanos , Pulmão/diagnóstico por imagem , Imageamento por Ressonância Magnética/métodos , Nanopartículas/química , Soroalbumina BovinaRESUMO
Patients with multiple myeloma, an incurable malignancy of plasma cells, frequently develop osteolytic bone lesions that severely impact quality of life and clinical outcomes. Eliglustat, a U.S. Food and Drug Administration-approved glucosylceramide synthase inhibitor, reduced osteoclast-driven bone loss in preclinical in vivo models of myeloma. In combination with zoledronic acid, a bisphosphonate that treats myeloma bone disease, eliglustat provided further protection from bone loss. Autophagic degradation of TRAF3, a key step for osteoclast differentiation, was inhibited by eliglustat as evidenced by TRAF3 lysosomal and cytoplasmic accumulation. Eliglustat blocked autophagy by altering glycosphingolipid composition whilst restoration of missing glycosphingolipids rescued autophagy markers and TRAF3 degradation thus restoring osteoclastogenesis in bone marrow cells from myeloma patients. This work delineates both the mechanism by which glucosylceramide synthase inhibition prevents autophagic degradation of TRAF3 to reduce osteoclastogenesis as well as highlighting the clinical translational potential of eliglustat for the treatment of myeloma bone disease.
Assuntos
Doenças Ósseas , Mieloma Múltiplo , Humanos , Mieloma Múltiplo/patologia , Fator 3 Associado a Receptor de TNF/metabolismo , Qualidade de Vida , Osteoclastos/metabolismo , Doenças Ósseas/tratamento farmacológico , Doenças Ósseas/metabolismo , Autofagia , Glicoesfingolipídeos/metabolismoRESUMO
It has been suggested that mitochondrial dysfunction and mtDNA variations may contribute to osteoarthritis (OA) pathogenesis. However, the causative link to support this claim is lacking. Here, we surgically-induced OA in conplastic mice in order to evaluate the functional consequences of mtDNA haplotypes in their joint degeneration. BL/6NZB strain was developed with C57BL/6JOlaHsd nuclear genome and NZB/OlaHsdmtDNA while BL/6C57, which is the original, was developed with C57BL/6JOlaHsd nuclear genome and C57/OlaHsdmtDNA for comparison. The surgical DMM OA model was induced in both strains. Their knees were processed and examined for histopathological changes. Cartilage expression of markers of autophagy, apoptosis, oxidative stress and senescence were also analyzed by immunohistochemistry. The joints of BL/6NZB mice that were operated presented more cellularity together with a reduced OARSI histopathology score, subchondral bone, menisci score and synovitis compared to those of BL/6C57 mice. This was accompanied with higher autophagy and a lower apoptosis in the cartilage of BL/6NZB mice that were operated. Therefore, the study demonstrates the functional impact of non-pathological variants of mtDNA on OA process using a surgically-induced OA model. Conplastic (BL/6NZB ) mice develop less severe OA compared to the BL/6C57original strain. These findings demonstrate that mitochondria and mtDNA are critical targets for potential novel therapeutic approaches to treat osteoarthritis.
Assuntos
DNA Mitocondrial , Osteoartrite/genética , Osteoartrite/fisiopatologia , Animais , Apoptose/genética , Autofagia , Cartilagem Articular/patologia , Modelos Animais de Doenças , Masculino , Meniscos Tibiais/fisiopatologia , Meniscos Tibiais/cirurgia , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Osteoartrite/etiologia , Sinovite/etiologia , Sinovite/genéticaRESUMO
Epitopes derived from mutated cancer proteins elicit strong antitumor T-cell responses that correlate with clinical efficacy in a proportion of patients. However, it remains unclear whether the subcellular localization of mutated proteins influences the efficiency of T-cell priming. To address this question, we compared the immunogenicity of NY-ESO-1 and OVA localized either in the cytosol or in mitochondria. We showed that tumors expressing mitochondrial-localized NY-ESO-1 and OVA proteins elicit significantdly higher frequencies of antigen-specific CD8+ T cells in vivo. We also demonstrated that this stronger immune response is dependent on the mitochondrial location of the antigenic proteins, which contributes to their higher steady-state amount, compared with cytosolic localized proteins. Consistent with these findings, we showed that injection of mitochondria purified from B16 melanoma cells can protect mice from a challenge with B16 cells, but not with irrelevant tumors. Finally, we extended these findings to cancer patients by demonstrating the presence of T-cell responses specific for mutated mitochondrial-localized proteins. These findings highlight the utility of prioritizing epitopes derived from mitochondrial-localized mutated proteins as targets for cancer vaccination strategies.
Assuntos
Antígenos de Neoplasias/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Vacinas Anticâncer/imunologia , Epitopos/imunologia , Proteínas Mitocondriais/imunologia , Neoplasias/imunologia , Animais , Linhagem Celular Tumoral , Modelos Animais de Doenças , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Proteínas Mitocondriais/metabolismo , Neoplasias/metabolismo , Neoplasias/terapiaRESUMO
Heteroplasmy, multiple variants of mitochondrial DNA (mtDNA) in the same cytoplasm, may be naturally generated by mutations but is counteracted by a genetic mtDNA bottleneck during oocyte development. Engineered heteroplasmic mice with nonpathological mtDNA variants reveal a nonrandom tissue-specific mtDNA segregation pattern, with few tissues that do not show segregation. The driving force for this dynamic complex pattern has remained unexplained for decades, challenging our understanding of this fundamental biological problem and hindering clinical planning for inherited diseases. Here, we demonstrate that the nonrandom mtDNA segregation is an intracellular process based on organelle selection. This cell type-specific decision arises jointly from the impact of mtDNA haplotypes on the oxidative phosphorylation (OXPHOS) system and the cell metabolic requirements and is strongly sensitive to the nuclear context and to environmental cues.
RESUMO
mtDNA is present in multiple copies in each cell derived from the expansions of those in the oocyte. Heteroplasmy, more than one mtDNA variant, may be generated by mutagenesis, paternal mtDNA leakage, and novel medical technologies aiming to prevent inheritance of mtDNA-linked diseases. Heteroplasmy phenotypic impact remains poorly understood. Mouse studies led to contradictory models of random drift or haplotype selection for mother-to-offspring transmission of mtDNA heteroplasmy. Here, we show that mtDNA heteroplasmy affects embryo metabolism, cell fitness, and induced pluripotent stem cell (iPSC) generation. Thus, genetic and pharmacological interventions affecting oxidative phosphorylation (OXPHOS) modify competition among mtDNA haplotypes during oocyte development and/or at early embryonic stages. We show that heteroplasmy behavior can fall on a spectrum from random drift to strong selection, depending on mito-nuclear interactions and metabolic factors. Understanding heteroplasmy dynamics and its mechanisms provide novel knowledge of a fundamental biological process and enhance our ability to mitigate risks in clinical applications affecting mtDNA transmission.
Assuntos
DNA Mitocondrial/genética , Desenvolvimento Embrionário/genética , Herança Materna/genética , Mitocôndrias/genética , Oogênese/genética , Animais , Linhagem Celular , Embrião de Mamíferos , Feminino , Fibroblastos , Haplótipos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , OócitosRESUMO
Post-translational modifications hugely increase the functional diversity of proteomes. Recent algorithms based on ultratolerant database searching are forging a path to unbiased analysis of peptide modifications by shotgun mass spectrometry. However, these approaches identify only one-half of the modified forms potentially detectable and do not map the modified residue. Moreover, tools for the quantitative analysis of peptide modifications are currently lacking. Here, we present a suite of algorithms that allows comprehensive identification of detectable modifications, pinpoints the modified residues, and enables their quantitative analysis through an integrated statistical model. These developments were used to characterize the impact of mitochondrial heteroplasmy on the proteome and on the modified peptidome in several tissues from 12-week-old mice. Our results reveal that heteroplasmy mainly affects cardiac tissue, inducing oxidative damage to proteins of the oxidative phosphorylation system, and provide a molecular mechanism explaining the structural and functional alterations produced in heart mitochondria.
Assuntos
Mitocôndrias Cardíacas/patologia , Miocárdio/metabolismo , Miocárdio/patologia , Estresse Oxidativo , Proteoma/metabolismo , Proteômica/métodos , Animais , Células HEK293 , Humanos , Masculino , Camundongos Endogâmicos C57BL , Mitocôndrias Cardíacas/metabolismo , Fosforilação Oxidativa , Peptídeos/metabolismo , Processamento de Proteína Pós-TraducionalRESUMO
Heart failure (HF) is a major health and economic burden in developed countries. It has been proposed that the pathogenesis of HF may involve the action of mitochondria. We evaluate three different mouse models of HF: tachycardiomyopathy, HF with preserved left ventricular (LV) ejection fraction (LVEF), and LV myocardial ischemia and hypertrophy. Regardless of whether LVEF is preserved, our results indicate that the three models share common features: an increase in mitochondrial reactive oxygen species followed by ultrastructural alterations in the mitochondrial cristae and loss of mitochondrial integrity that lead to cardiomyocyte death. We show that the ablation of the mitochondrial protease OMA1 averts cardiomyocyte death in all three murine HF models, and thus loss of OMA1 plays a direct role in cardiomyocyte protection. This finding identifies OMA1 as a potential target for preventing the progression of myocardial damage in HF associated with a variety of etiologies.
Assuntos
Insuficiência Cardíaca/metabolismo , Metaloproteases/metabolismo , Proteínas Mitocondriais/metabolismo , Animais , Insuficiência Cardíaca/genética , Masculino , Metaloproteases/genética , Camundongos , Mitocôndrias/metabolismo , Proteínas Mitocondriais/genética , Miócitos Cardíacos/metabolismo , Espécies Reativas de Oxigênio/metabolismoRESUMO
Increasing the thermogenic capacity of adipose tissue to enhance organismal energy expenditure is considered a promising therapeutic strategy to combat obesity. Here, we report that expression of the p38 MAPK activator MKK6 is elevated in white adipose tissue of obese individuals. Using knockout animals and shRNA, we show that Mkk6 deletion increases energy expenditure and thermogenic capacity of white adipose tissue, protecting mice against diet-induced obesity and the development of diabetes. Deletion of Mkk6 increases T3-stimulated UCP1 expression in adipocytes, thereby increasing their thermogenic capacity. Mechanistically, we demonstrate that, in white adipose tissue, p38 is activated by an alternative pathway involving AMPK, TAK, and TAB. Our results identify MKK6 in adipocytes as a potential therapeutic target to reduce obesity.Brown and beige adipose tissues dissipate heat via uncoupling protein 1 (UCP1). Here the authors show that the stress activated kinase MKK6 acts as a repressor of UCP1 expression, suggesting that its inhibition promotes adipose tissue browning and increases organismal energy expenditure.