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In this review we examine the functionally diverse ATPase associated with various cellular activities (AAA-ATPase), valosin-containing protein (VCP/p97), its molecular functions, the mutational landscape of VCP and the phenotypic manifestation of VCP disease. VCP is crucial to a multitude of cellular functions including protein quality control, endoplasmic reticulum-associated degradation (ERAD), autophagy, mitophagy, lysophagy, stress granule formation and clearance, DNA replication and mitosis, DNA damage response including nucleotide excision repair, ATM- and ATR-mediated damage response, homologous repair and non-homologous end joining. VCP variants cause multisystem proteinopathy, and pathology can arise in several tissue types such as skeletal muscle, bone, brain, motor neurons, sensory neurons and possibly cardiac muscle, with the disease course being challenging to predict.
Assuntos
Fenótipo , Proteína com Valosina , Proteína com Valosina/metabolismo , Proteína com Valosina/genética , Humanos , Animais , Mutação , Autofagia/genética , Reparo do DNARESUMO
Mitochondria are pleiotropic organelles central to an array of cellular pathways including metabolism, signal transduction, and programmed cell death. Mitochondria are also key drivers of mammalian immune responses, functioning as scaffolds for innate immune signaling, governing metabolic switches required for immune cell activation, and releasing agonists that promote inflammation. Mitochondrial DNA (mtDNA) is a potent immunostimulatory agonist, triggering pro-inflammatory and type I interferon responses in a host of mammalian cell types. Here we review recent advances in how mtDNA is detected by nucleic acid sensors of the innate immune system upon release into the cytoplasm and extracellular space. We also discuss how the interplay between mtDNA release and sensing impacts cellular innate immune endpoints relevant to health and disease.
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DNA Mitocondrial , Imunidade Inata , Mitocôndrias , Transdução de Sinais , Humanos , DNA Mitocondrial/genética , DNA Mitocondrial/imunologia , Mitocôndrias/metabolismo , Mitocôndrias/imunologia , Mitocôndrias/genética , Animais , Transdução de Sinais/imunologia , Interferon Tipo I/imunologia , Interferon Tipo I/metabolismo , Interferon Tipo I/genética , Inflamação/imunologia , Inflamação/genéticaRESUMO
Aims: Structural analogues of bisphenol A (BPA), including bisphenol S (BPS) and bisphenol F (BPF), are emerging environmental toxicants as their presence in the environment is rising since new regulatory restrictions were placed on BPA-containing infant products. The adipogenesis-enhancing effect of bisphenols may explain the link between human exposure and metabolic disease; however, underlying molecular pathways remain unresolved. Results: Exposure to BPS, BPF, BPA, or reactive oxygen species (ROS) generators enhanced lipid droplet formation and expression of adipogenic markers after induction of differentiation in adipose-derived progenitors isolated from mice. RNAseq analysis in BPS-exposed progenitors revealed modulation in pathways regulating adipogenesis and responses to oxidative stress. ROS were higher in bisphenol-exposed cells, while cotreatment with antioxidants attenuated adipogenesis and abolished the effect of BPS. There was a loss of mitochondrial membrane potential in BPS-exposed cells and mitochondria-derived ROS contributed to the potentiation of adipogenesis by BPS and its analogues. Male mice exposed to BPS during gestation had higher whole-body adiposity, as measured by time domain nuclear magnetic resonance, while postnatal exposure had no impact on adiposity in either sex. Innovation: These findings support existing evidence showing a role for ROS in regulating adipocyte differentiation and are the first to highlight ROS as a unifying mechanism that explains the proadipogenic properties of BPA and its structural analogues. Conclusion: ROS act as signaling molecules in the regulation of adipocyte differentiation and mediate bisphenol-induced potentiation of adipogenesis. Antioxid. Redox Signal. 40, 1-15.
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Adipogenia , Compostos Benzidrílicos , Fenóis , Sulfonas , Humanos , Masculino , Camundongos , Animais , Espécies Reativas de Oxigênio , Compostos Benzidrílicos/farmacologiaRESUMO
Mitochondrial quality control (MQC) mechanisms are required to maintain a functional proteome, which enables mitochondria to perform a myriad of important cellular functions from oxidative phosphorylation to numerous other metabolic pathways. Mitochondrial protein homeostasis begins with the import of over 1000 nuclear-encoded mitochondrial proteins and the synthesis of 13 mitochondrial DNA-encoded proteins. A network of chaperones and proteases helps to fold new proteins and degrade unnecessary, damaged, or misfolded proteins, whereas more extensive damage can be removed by mitochondrial-derived vesicles (MDVs) or mitochondrial autophagy (mitophagy). Here, focusing on mechanisms in mammalian cells, we review the importance of mitochondrial protein import as a sentinel of mitochondrial function that activates multiple MQC mechanisms when impaired.
Assuntos
Autofagia , Mitocôndrias , Animais , Humanos , Mitocôndrias/metabolismo , Mitofagia , Resposta a Proteínas não Dobradas , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Mamíferos/metabolismoRESUMO
Gut bacteria provide benefits to the host and have been implicated in inflammatory bowel disease (IBD), where adherent-invasive E. coli (AIEC) pathobionts (e.g., strain LF82) are associated with Crohn's disease. E. coli-LF82 causes fragmentation of the epithelial mitochondrial network, leading to increased epithelial permeability. We hypothesized that butyrate would limit the epithelial mitochondrial disruption caused by E. coli-LF82. Human colonic organoids and the T84 epithelial cell line infected with E. coli-LF82 (MOI = 100, 4 h) showed a significant increase in mitochondrial network fission that was reduced by butyrate (10 mM) co-treatment. Butyrate reduced the loss of mitochondrial membrane potential caused by E. coli-LF82 and increased expression of PGC-1α mRNA, the master regulator of mitochondrial biogenesis. Metabolomics revealed that butyrate significantly altered E. coli-LF82 central carbon metabolism leading to diminished glucose uptake and increased succinate secretion. Correlating with preservation of mitochondrial network form/function, butyrate reduced E. coli-LF82 transcytosis across T84-cell monolayers. The use of the G-protein inhibitor, pertussis toxin, implicated GPCR signaling as critical to the effect of butyrate, and the free fatty acid receptor three (FFAR3, GPR41) agonist, AR420626, reproduced butyrate's effect in terms of ameliorating the loss of barrier function and reducing the mitochondrial fragmentation observed in E. coli-LF82 infected T84-cells and organoids. These data indicate that butyrate helps maintain epithelial mitochondrial form/function when challenged by E. coli-LF82 and that this occurs, at least in part, via FFAR3. Thus, loss of butyrate-producing bacteria in IBD in the context of pathobionts would contribute to loss of epithelial mitochondrial and barrier functions that could evoke disease and/or exaggerate a low-grade inflammation.
Assuntos
Doença de Crohn , Infecções por Escherichia coli , Microbioma Gastrointestinal , Humanos , Escherichia coli/genética , Infecções por Escherichia coli/microbiologia , Mucosa Intestinal/microbiologia , Ácidos Graxos não Esterificados/metabolismo , Butiratos/farmacologia , Butiratos/metabolismo , Doença de Crohn/microbiologia , Aderência Bacteriana/genéticaRESUMO
Prolonged severe hypoxia follows brief seizures and represents a mechanism underlying several negative postictal manifestations without interventions. Approximately 50% of the postictal hypoxia phenomenon can be accounted for by arteriole vasoconstriction. What accounts for the rest of the drop in unbound oxygen is unclear. Here, we determined the effect of pharmacological modulation of mitochondrial function on tissue oxygenation in the hippocampus of rats after repeatedly evoked seizures. Rats were treated with mitochondrial uncoupler 2,4 dinitrophenol (DNP) or antioxidants. Oxygen profiles were recorded using a chronically implanted oxygen-sensing probe, before, during, and after seizure induction. Mitochondrial function and redox tone were measured using in vitro mitochondrial assays and immunohistochemistry. Postictal cognitive impairment was assessed using the novel object recognition task. Mild mitochondrial uncoupling by DNP raised hippocampal oxygen tension and ameliorated postictal hypoxia. Chronic DNP also lowered mitochondrial oxygen-derived reactive species and oxidative stress in the hippocampus during postictal hypoxia. Uncoupling the mitochondria exerts therapeutic benefits on postictal cognitive dysfunction. Finally, antioxidants do not affect postictal hypoxia, but protect the brain from associated cognitive deficits. We provided evidence for a metabolic component of the prolonged oxygen deprivation that follow seizures and its pathological sequelae. Furthermore, we identified a molecular underpinning of this metabolic component, which involves excessive oxygen conversion into reactive species. Mild mitochondrial uncoupling may be a potential therapeutic strategy to treat the postictal state where seizure control is absent or poor.
Assuntos
Antioxidantes , Hipóxia , Ratos , Animais , Espécies Reativas de Oxigênio/metabolismo , Antioxidantes/farmacologia , Hipóxia/metabolismo , Oxigênio/metabolismo , Mitocôndrias , Convulsões/metabolismo , Desacopladores/metabolismo , Desacopladores/farmacologiaRESUMO
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.
Assuntos
Cardiotoxicidade , DNA Mitocondrial , Animais , Camundongos , DNA Mitocondrial/metabolismo , Imunidade Inata , Interferons/metabolismo , Nucleotidiltransferases/genética , Nucleotidiltransferases/metabolismo , FosforilaçãoRESUMO
The recognition that cytosolic mitochondrial DNA (mtDNA) activates cyclic GMP-AMP synthase-stimulator of interferon genes (cGAS-STING) innate immune signaling has unlocked novel disease mechanisms. Here, an uncharacterized variant predicted to affect TOP1MT function, P193L, was discovered in a family with multiple early onset autoimmune diseases, including Systemic Lupus Erythematosus (SLE). Although there was no previous genetic association between TOP1MT and autoimmune disease, the role of TOP1MT as a regulator of mtDNA led us to investigate whether TOP1MT could mediate the release of mtDNA to the cytosol, where it could then activate the cGAS-STING innate immune pathway known to be activated in SLE and other autoimmune diseases. Through analysis of cells with reduced TOP1MT expression, we show that loss of TOP1MT results in release of mtDNA to the cytosol, which activates the cGAS-STING pathway. We also characterized the P193L variant for its ability to rescue several TOP1MT functions when expressed in TOP1MT knockout cells. We show that the P193L variant is not fully functional, as its re-expression at high levels was unable to rescue mitochondrial respiration deficits, and only showed partial rescue for other functions, including repletion of mtDNA replication following depletion, nucleoid size, steady state mtDNA transcripts levels and mitochondrial morphology. Additionally, expression of P193L at endogenous levels was unable to rescue mtDNA release-mediated cGAS-STING signaling. Overall, we report a link between TOP1MT and mtDNA release leading to cGAS-STING activation. Moreover, we show that the P193L variant has partial loss of function that may contribute to autoimmune disease susceptibility via cGAS-STING mediated activation of the innate immune system.
Assuntos
Doenças Autoimunes , Lúpus Eritematoso Sistêmico , Humanos , DNA Mitocondrial/genética , Imunidade Inata/genética , Interferons , Nucleotidiltransferases/genética , Nucleotidiltransferases/metabolismoRESUMO
Mitochondrial dysfunction as defined by transcriptomic and proteomic analysis of biopsies or ultra-structure in transmission electron microscopy occurs in inflammatory bowel disease (IBD); however, mitochondrial dynamics in IBD have received minimal attention, with most investigations relying on cell-based in vitro models. We build on these studies by adapting the epithelial cell immunofluorescence workflow to imaging mitochondrial networks in normal and inflamed colonic tissue (i.e., murine di-nitrobenzene sulphonic acid (DNBS)-induced colitis, human ulcerative colitis). Using antibodies directed to TOMM20 (translocase of outer mitochondrial membrane 20) and cytochrome-C, we have translated the cell-based protocol for high-fidelity imaging to examine epithelial mitochondria networks in intact intestine. In epithelia of non-inflamed small or large intestinal tissue, the mitochondrial networks were dense and compact. This pattern was more pronounced in the basal region of the cell compared to that between the nucleus and apical surface facing the gut lumen. In comparison, mitochondrial networks in inflamed tissue displayed substantial loss of TOMM20+ staining. The remaining networks were less dense and fragmented, and contained isolated spherical mitochondrial fragments. The degree of mitochondrial network fragmentation mirrored the severity of inflammation, as assessed by blinded semi-quantitative scoring. As an indication of poor cell 'health' or viability, cytosolic cytochrome-C was observed in enterocytes with highly fragmented mitochondria. Thus, high-resolution and detailed visualization of mitochondrial networks in tissue is a feasible and valuable approach to assess disease, suited to characterizing mitochondrial abnormalities in tissue. We speculate that drugs that maintain a functional remodelling mitochondrial network and limit excess fragmentation could be a valuable addition to current therapies for IBD.
Assuntos
Citocromos c , Doenças Inflamatórias Intestinais , Humanos , Camundongos , Animais , Citocromos c/metabolismo , Proteômica , Colo/metabolismo , Colo/patologia , Doenças Inflamatórias Intestinais/metabolismo , Doenças Inflamatórias Intestinais/patologia , Proteínas de Transporte , Mitocôndrias/metabolismoRESUMO
TOP1MT encodes a mitochondrial topoisomerase that is important for mtDNA regulation and is involved in mitochondrial replication, transcription, and translation. Two variants predicted to affect TOP1MT function (V1 - R198C and V2 - V338L) were identified by exome sequencing of a newborn with hypertrophic cardiomyopathy. As no pathogenic TOP1MT variants had been confirmed previously, we characterized these variants for their ability to rescue several TOP1MT functions in KO cells. Consistent with these TOP1MT variants contributing to the patient phenotype, our comprehensive characterization suggests that both variants had impaired activity. Critically, we determined neither variant was able to restore steady state levels of mitochondrial-encoded proteins nor to rescue oxidative phosphorylation when re-expressed in TOP1MT KO cells. However, we found the two variants behaved differently in some respects; while the V1 variant was more efficient in restoring transcript levels, the V2 variant showed better rescue of mtDNA copy number and replication. These findings suggest that the different TOP1MT variants affect distinct TOP1MT functions. Altogether, these findings begin to provide insight into the many roles that TOP1MT plays in the maintenance and expression of the mitochondrial genome and how impairments in this important protein may lead to human pathology.
Assuntos
Cardiomiopatia Hipertrófica , DNA Topoisomerases Tipo I , Genoma Mitocondrial , Mitocôndrias , Humanos , Recém-Nascido , Cardiomiopatia Hipertrófica/genética , DNA Topoisomerases Tipo I/genética , DNA Topoisomerases Tipo I/metabolismo , DNA Mitocondrial/metabolismo , Variação Genética , Mitocôndrias/enzimologia , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismoRESUMO
Intestinal homeostasis is highly dependent on optimal epithelial barrier function and permeability. Intestinal epithelial cells (IEC) regulate these properties acting as cellular gatekeepers by selectively absorbing nutrients and controlling the passage of luminal bacteria. These functions are energy demanding processes that are presumably met through mitochondrial-based processes. Routine methods for examining IEC mitochondrial function remain sparse, hence, our objective is to present standardized methods for quantifying mitochondrial energetics in an immortalized IEC line. Employing the murine IEC4.1 cell line, we present adapted methods and protocols to examine mitochondrial function using two well-known platforms: the Seahorse Extracellular Flux Analyzer and Oxygraph-2 k. To demonstrate the applicability of these protocols and instruments, IEC were treated with and without the murine colitogenic agent, dextran sulfate sodium (DSS, 2% w/v). Profound impairments with DSS treatment were found with both platforms, however, the Oxygraph-2 k allowed greater resolution of affected pathways including short-chain fatty acid metabolism. Mitochondrial functional analysis is a novel tool to explore the relationship between IEC energetics and functional consequences within the contexts of health and disease. The outlined methods offer an introductory starting point for such assessment and provide the investigator with insights into platform-specific capabilities.
Assuntos
Colite , Mucosa Intestinal , Animais , Colite/induzido quimicamente , Colite/metabolismo , Sulfato de Dextrana/toxicidade , Metabolismo Energético , Células Epiteliais/metabolismo , Mucosa Intestinal/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/metabolismoRESUMO
The Mitofusin 2 protein (MFN2), encoded by the MFN2 gene, was first described for its role in mediating mitochondrial fusion. However, MFN2 is now recognized to play additional roles in mitochondrial autophagy (mitophagy), mitochondrial motility, lipid transfer, and as a tether to other organelles including the endoplasmic reticulum (ER) and lipid droplets. The tethering role of MFN2 is an important mediator of mitochondrial-ER contact sites (MERCs), which themselves have many important functions that regulate mitochondria, including calcium homeostasis and lipid metabolism. Exemplifying the importance of MFN2, pathogenic variants in MFN2 are established to cause the peripheral neuropathy Charcot-Marie-Tooth Disease Subtype 2A (CMT2A). However, the mechanistic basis for disease is not clear. Moreover, additional pathogenic phenotypes such as lipomatosis, distal myopathy, optic atrophy, and hearing loss, can also sometimes be present in patients with CMT2A. Given these variable patient phenotypes, and the many cellular roles played by MFN2, the mechanistic underpinnings of the cellular impairments by which MFN2 dysfunction leads to disease are likely to be complex. Here, we will review what is known about the various functions of MFN2 that are impaired by pathogenic variants causing CMT2A, with a specific emphasis on the ties between MFN2 variants and MERCs.
RESUMO
Human mitochondrial disorders impact tissues with high energetic demands and can be associated with cardiac muscle disease (cardiomyopathy) and early mortality. However, the mechanistic link between mitochondrial disease and the development of cardiomyopathy is frequently unclear. In addition, there is often marked phenotypic heterogeneity between patients, even between those with the same genetic variant, which is also not well understood. Several of the mitochondrial cardiomyopathies are related to defects in the maintenance of mitochondrial protein homeostasis, or proteostasis. This essential process involves the importing, sorting, folding and degradation of preproteins into fully functional mature structures inside mitochondria. Disrupted mitochondrial proteostasis interferes with mitochondrial energetics and ATP production, which can directly impact cardiac function. An inability to maintain proteostasis can result in mitochondrial dysfunction and subsequent mitophagy or even apoptosis. We review the known mitochondrial diseases that have been associated with cardiomyopathy and which arise from mutations in genes that are important for mitochondrial proteostasis. Genes discussed include DnaJ heat shock protein family member C19 (DNAJC19), mitochondrial import inner membrane translocase subunit TIM16 (MAGMAS), translocase of the inner mitochondrial membrane 50 (TIMM50), mitochondrial intermediate peptidase (MIPEP), X-prolyl-aminopeptidase 3 (XPNPEP3), HtraA serine peptidase 2 (HTRA2), caseinolytic mitochondrial peptidase chaperone subunit B (CLPB) and heat shock 60-kD protein 1 (HSPD1). The identification and description of disorders with a shared mechanism of disease may provide further insights into the disease process and assist with the identification of potential therapeutics.
Assuntos
Cardiomiopatias , Proteínas Mitocondriais , Cardiomiopatias/genética , Cardiomiopatias/metabolismo , Proteínas de Choque Térmico HSP40/metabolismo , Homeostase , Humanos , Mitocôndrias/metabolismo , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Peptídeo Hidrolases/metabolismo , Transporte Proteico , ProteostaseRESUMO
The dynamic nature of mitochondria, which can fuse, divide and move throughout the cell, allows these critical organelles to adapt their function in response to cellular demands, and is also important for regulating mitochondrial DNA (mtDNA). While it is established that impairments in mitochondrial fusion and fission impact the mitochondrial genome and can lead to mtDNA depletion, abnormal nucleoid organization or accumulation of deletions, it is not entirely clear how or why remodeling mitochondrial network morphology affects mtDNA. Here, we focus on recent advances in our understanding of how mitochondrial dynamics contribute to the regulation of mtDNA and discuss links to human disease.
Assuntos
Genoma Mitocondrial , Dinâmica Mitocondrial , DNA Mitocondrial/genética , Humanos , Mitocôndrias/genética , Dinâmica Mitocondrial/genéticaRESUMO
The dynamic processes of mitochondrial fusion and fission determine the shape of mitochondria, which can range from individual fragments to a hyperfused network, and influence mitochondrial function. Changes in mitochondrial shape can occur rapidly, allowing mitochondria to adapt to specific cues and changing cellular demands. Here, we will review what is known about how key proteins required for mitochondrial fusion and fission are regulated by their acetylation status, with acetylation promoting fission and deacetylation enhancing fusion. In particular, we will examine the roles of NAD+ dependant sirtuin deacetylases, which mediate mitochondrial acetylation, and how this post-translational modification provides an exquisite regulatory mechanism to co-ordinate mitochondrial function with metabolic demands of the cell.
Assuntos
Mitocôndrias/fisiologia , Dinâmica Mitocondrial , Proteínas/metabolismo , AcetilaçãoRESUMO
Mitochondrial DNA (mtDNA) maintenance is essential to sustain a functionally healthy population of mitochondria within cells. Proper mtDNA replication and distribution within mitochondrial networks are essential to maintain mitochondrial homeostasis. However, the fundamental basis of mtDNA segregation and distribution within mitochondrial networks is still unclear. To address these questions, we developed an algorithm, Mitomate tracker to unravel the global distribution of nucleoids within mitochondria. Using this tool, we decipher the semi-regular spacing of nucleoids across mitochondrial networks. Furthermore, we show that mitochondrial fission actively regulates mtDNA distribution by controlling the distribution of nucleoids within mitochondrial networks. Specifically, we found that primary cells bearing disease-associated mutations in the fission proteins DRP1 and MYH14 show altered nucleoid distribution, and acute enrichment of enlarged nucleoids near the nucleus. Further analysis suggests that the altered nucleoid distribution observed in the fission mutants is the result of both changes in network structure and nucleoid density. Thus, our study provides novel insights into the role of mitochondria fission in nucleoid distribution and the understanding of diseases caused by fission defects.
Assuntos
Núcleo Celular/metabolismo , DNA Mitocondrial/metabolismo , Dinaminas/metabolismo , Homeostase , Mitocôndrias/metabolismo , Dinâmica Mitocondrial , Cadeias Pesadas de Miosina/metabolismo , Miosina Tipo II/metabolismo , Núcleo Celular/genética , Replicação do DNA , DNA Mitocondrial/genética , Dinaminas/genética , Humanos , Mitocôndrias/genética , Cadeias Pesadas de Miosina/genética , Miosina Tipo II/genéticaRESUMO
Spermatogonia are stem and progenitor cells responsible for maintaining mammalian spermatogenesis. Preserving the balance between self-renewal of spermatogonial stem cells (SSCs) and differentiation is critical for spermatogenesis and fertility. Ubiquitin carboxy-terminal hydrolase-L1 (UCH-L1) is highly expressed in spermatogonia of many species; however, its functional role has not been identified. Here, we aimed to understand the role of UCH-L1 in murine spermatogonia using a Uch-l1-/- mouse model. We confirmed that UCH-L1 is expressed in undifferentiated and early-differentiating spermatogonia in the post-natal mammalian testis. The Uch-l1-/- mice showed reduced testis weight and progressive degeneration of seminiferous tubules. Single-cell transcriptome analysis detected a dysregulated metabolic profile in spermatogonia of Uch-l1-/- compared to wild-type mice. Furthermore, cultured Uch-l1-/- SSCs had decreased capacity in regenerating full spermatogenesis after transplantation in vivo and accelerated oxidative phosphorylation (OXPHOS) during maintenance in vitro. Together, these results indicate that the absence of UCH-L1 impacts the maintenance of SSC homeostasis and metabolism and impacts the differentiation competence. Metabolic perturbations associated with loss of UCH-L1 appear to underlie a reduced capacity for supporting spermatogenesis and fertility with age. This work is one step further in understanding the complex regulatory circuits underlying SSC function.
Assuntos
Diferenciação Celular , Regulação da Expressão Gênica no Desenvolvimento , Mitocôndrias/patologia , Espermatogênese , Espermatogônias/patologia , Células-Tronco/patologia , Ubiquitina Tiolesterase/fisiologia , Animais , Células Cultivadas , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mitocôndrias/metabolismo , Análise de Sequência de RNA , Análise de Célula Única , Espermatogônias/metabolismo , Células-Tronco/metabolismoRESUMO
Mitochondria are dynamic organelles capable of fusing, dividing, and moving about the cell. These properties are especially important in neurons, which in addition to high energy demand, have unique morphological properties with long axons. Notably, mitochondrial dysfunction causes a variety of neurological disorders including peripheral neuropathy, which is linked to impaired mitochondrial dynamics. Nonetheless, exactly why peripheral neurons are especially sensitive to impaired mitochondrial dynamics remains somewhat enigmatic. Although the prevailing view is that longer peripheral nerves are more sensitive to the loss of mitochondrial motility, this explanation is insufficient. Here, we review pathogenic variants in proteins mediating mitochondrial fusion, fission and transport that cause peripheral neuropathy. In addition to highlighting other dynamic processes that are impacted in peripheral neuropathies, we focus on impaired mitochondrial quality control as a potential unifying theme for why mitochondrial dysfunction and impairments in mitochondrial dynamics in particular cause peripheral neuropathy.
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Phosphatidylethanolamine (PE) made in mitochondria has long been recognized as an important precursor for phosphatidylcholine production that occurs in the endoplasmic reticulum (ER). Recently, the strict mitochondrial localization of the enzyme that makes PE in the mitochondrion, phosphatidylserine decarboxylase 1 (Psd1), was questioned. Since a dual localization of Psd1 to the ER would have far-reaching implications, we initiated our study to independently re-assess the subcellular distribution of Psd1. Our results support the unavoidable conclusion that the vast majority, if not all, of functional Psd1 resides in the mitochondrion. Through our efforts, we discovered that mutant forms of Psd1 that impair a self-processing step needed for it to become functional are dually localized to the ER when expressed in a PE-limiting environment. We conclude that severely impaired cellular PE metabolism provokes an ER-assisted adaptive response that is capable of identifying and resolving nonfunctional mitochondrial precursors.
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BACKGROUND & AIMS: Adherent-invasive Escherichia coli are implicated in inflammatory bowel disease, and mitochondrial dysfunction has been observed in biopsy specimens from patients with inflammatory bowel disease. As a novel aspect of adherent-invasive E coli-epithelial interaction, we hypothesized that E coli (strain LF82) would elicit substantial disruption of epithelial mitochondrial form and function. METHODS: Monolayers of human colon-derived epithelial cell lines were exposed to E coli-LF82 or commensal E coli and RNA sequence analysis, mitochondrial function (adenosine triphosphate synthesis) and dynamics (mitochondrial network imaging, immunoblotting for fission and fusion proteins), and epithelial permeability (transepithelial resistance, flux of fluorescein isothiocyanate-dextran and bacteria) were assessed. RESULTS: E coli-LF82 significantly affected epithelial expression of â¼8600 genes, many relating to mitochondrial function. E coli-LF82-infected epithelia showed swollen mitochondria, reduced mitochondrial membrane potential and adenosine triphosphate, and fragmentation of the mitochondrial network: events not observed with dead E coli-LF82, medium from bacterial cultures, or control E coli. Treatment with Mitochondrial Division Inhibitor 1 (Mdivi1, inhibits dynamin-related peptide 1, guanosine triphosphatase principally responsible for mitochondrial fission) or P110 (prevents dynamin-related peptide 1 binding to mitochondrial fission 1 protein) partially reduced E coli-LF82-induced mitochondrial fragmentation in the short term. E coli-LF82-infected epithelia showed loss of the long isoform of optic atrophy factor 1, which mediates mitochondrial fusion. Mitochondrial Division Inhibitor 1 reduced the magnitude of E coli-LF82-induced increased transepithelial flux of fluorescein isothiocyanate dextran. By 8 hours after infection, increased cytosolic cytochrome C and DNA fragmentation were apparent without evidence of caspase-3 or apoptosis inducing factor activation. CONCLUSIONS: Epithelial mitochondrial fragmentation caused by E coli-LF82 could be targeted to maintain cellular homeostasis and mitigate infection-induced loss of epithelial barrier function. Data have been deposited in NCBI's Gene Expression Omnibus and are accessible through GEO series accession numbers GSE154121 and GSE154122 (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE154121).