scholarly journals Nix induced mitochondrial fission, mitophagy, and myocyte insulin resistance are abrogated by PKA phosphorylation

2019 ◽  
Author(s):  
Simone Cristina da Silva Rosa ◽  
Matthew D. Martens ◽  
Jared T. Field ◽  
Lucas Nguyen ◽  
Stephanie M. Kereliuk ◽  
...  
Keyword(s):  
Gene Expression ◽  
Insulin Resistance ◽  
Mitochondrial Dynamics ◽  
Mitochondrial Fission ◽  
Fatty Acyl ◽  
Loss Of Function ◽  
Expression Array ◽  
Calcium Dependent ◽  
Human Myotubes ◽  
Phosphatidic Acids

AbstractLipotoxicity is a form of cellular stress caused by the accumulation of lipids resulting in mitochondrial dysfunction and insulin resistance in muscle. Previously, we demonstrated that the mitophagy receptor Nix is responsive to lipotoxicity and accumulates in response to diacylglycerols induced by high-fat (HF) feeding. In addition, previous studies have implicated autophagy and mitophagy in muscle insulin sensitivity. To provide a better understanding of these observations, we undertook gene expression array and shot-gun metabolomics studies in soleus muscle from rodents on an HF diet. Interestingly, we observed a modest reduction in several autophagy-related genes including Beclin-1, ATG3, and -5. Moreover, we observed alterations in the fatty acyl composition of cardiolipins and phosphatidic acids. Given the previously reported roles of these phospholipids and Nix in mitochondrial dynamics, we investigated aberrant mitochondrial fission and turn-over as a mechanism of myocyte insulin resistance. In a series of gain-of-function and loss-of-function experiments in rodent and human myotubes, we demonstrate that Nix accumulation triggers mitochondrial depolarization, fragmentation, calcium-dependent activation of DRP1, and mitophagy. In addition, Nix-induced mitochondrial fission leads to myotube insulin resistance through activation of mTOR-p70S6 kinase inhibition of IRS1, which is contingent on phosphatidic acids and Rheb. Finally, we demonstrate that Nix-induced mitophagy and insulin resistance can be reversed by direct phosphorylation of Nix by PKA, leading to the translocation of Nix from the mitochondria and sarcoplasmic reticulum to the cytosol. These findings provide insight into the role of Nix-induced mitophagy and myocyte insulin resistance during an overfed state when overall autophagy-related gene expression is reduced. Furthermore, our data suggests a mechanism by which exercise or pharmacological activation of PKA may overcome myocyte insulin resistance.Graphical Abstract

scholarly journals Drp1-mediated mitochondrial fission regulates calcium and F-actin dynamics during wound healing

2019 ◽  
Author(s):  
Susana Ponte ◽  
Lara Carvalho ◽  
Maria Gagliardi ◽  
Isabel Campos ◽  
Paulo J. Oliveira ◽  
...  
Keyword(s):  
Wound Healing ◽  
Mitochondrial Dynamics ◽  
Mitochondrial Fission ◽  
Actin Dynamics ◽  
Mitochondrial Calcium ◽  
Loss Of Function ◽  
Wound Edge ◽  
Epithelial Repair ◽  
Healing Impairment

AbstractMitochondria adapt to cellular needs by changes in morphology through fusion and fission events, referred to as mitochondrial dynamics. Mitochondrial function and morphology are intimately connected and the dysregulation of mitochondrial dynamics is linked to several human diseases. In this work, we investigated the role of mitochondrial dynamics in wound healing in the Drosophila embryonic epidermis. Mutants for mitochondrial fusion and fission proteins fail to close their wounds, indicating that the regulation of mitochondrial dynamics is required for wound healing. By live-imaging, we found that loss of function of the mitochondrial fission protein Dynamin-related protein 1 (Drp1) compromises the increase of cytosolic and mitochondrial calcium upon wounding and leads to F-actin defects at the wound edge, culminating in wound healing impairment. Our results highlight a new role for mitochondrial dynamics in the regulation of calcium and F-actin during epithelial repair.SummaryWe show that mitochondrial dynamics proteins are required for epithelial repair. In particular, Drp1 loss-of-function leads to defects in the dynamics of cytosolic and mitochondrial calcium and F-actin upon wounding.

scholarly journals A novel role for kynurenine 3-monooxygenase in mitochondrial dynamics

PLoS Genetics ◽  
2020 ◽  
Vol 16 (11) ◽  
pp. e1009129
Author(s):  
Daniel C. Maddison ◽  
Mónica Alfonso-Núñez ◽  
Aisha M. Swaih ◽  
Carlo Breda ◽  
Susanna Campesan ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Translational Regulation ◽  
Metabolic Flux ◽  
Mitochondrial Dynamics ◽  
Mitochondrial Fission ◽  
Kynurenine Pathway ◽  
Loss Of Function ◽  
Disease Associated Genes

The enzyme kynurenine 3-monooxygenase (KMO) operates at a critical branch-point in the kynurenine pathway (KP), the major route of tryptophan metabolism. As the KP has been implicated in the pathogenesis of several human diseases, KMO and other enzymes that control metabolic flux through the pathway are potential therapeutic targets for these disorders. While KMO is localized to the outer mitochondrial membrane in eukaryotic organisms, no mitochondrial role for KMO has been described. In this study, KMO deficient Drosophila melanogaster were investigated for mitochondrial phenotypes in vitro and in vivo. We find that a loss of function allele or RNAi knockdown of the Drosophila KMO ortholog (cinnabar) causes a range of morphological and functional alterations to mitochondria, which are independent of changes to levels of KP metabolites. Notably, cinnabar genetically interacts with the Parkinson’s disease associated genes Pink1 and parkin, as well as the mitochondrial fission gene Drp1, implicating KMO in mitochondrial dynamics and mitophagy, mechanisms which govern the maintenance of a healthy mitochondrial network. Overexpression of human KMO in mammalian cells finds that KMO plays a role in the post-translational regulation of DRP1. These findings reveal a novel mitochondrial role for KMO, independent from its enzymatic role in the kynurenine pathway.

scholarly journals Mitochondrial dynamic modulation exerts cardiometabolic protection in obese insulin-resistant rats

2019 ◽  
Vol 133 (24) ◽  
pp. 2431-2447 ◽  
Author(s):  
Chayodom Maneechote ◽  
Siripong Palee ◽  
Nattayaporn Apaijai ◽  
Sasiwan Kerdphoo ◽  
Thidarat Jaiwongkam ◽  
...  
Keyword(s):  
Insulin Resistance ◽  
Cardiac Function ◽  
Mitochondrial Dynamics ◽  
Combined Therapy ◽  
Mitochondrial Fission ◽  
Combined Treatment ◽  
Myocardial Cell ◽  
Normal Diet ◽  
Mitochondrial Dynamic ◽  
Insulin Resistant

Abstract Obese insulin resistance impairs cardiac mitochondrial dynamics by increasing mitochondrial fission and decreasing mitochondrial fusion, leading to mitochondrial damage, myocardial cell death and cardiac dysfunction. Therefore, inhibiting fission and promoting fusion could provide cardioprotection in this pre-diabetic condition. We investigated the combined effects of the mitochondrial fission inhibitor (Mdivi1) and fusion promoter (M1) on cardiac function in obese insulin-resistant rats. We hypothesized that Mdivi1 and M1 protect heart against obese insulin-resistant condition, but also there will be greater improvement using Mdivi1 and M1 as a combined treatment. Wistar rats (n=56, male) were randomly assigned to a high-fat diet (HFD) and normal diet (ND) fed groups. After feeding with either ND or HFD for 12 weeks, rats in each dietary group were divided into groups to receive either the vehicle, Mdivi1 (1.2 mg/kg, i.p.), M1 (2 mg/kg, i.p.) or combined treatment for 14 days. The cardiac function, cardiac mitochondrial function, metabolic and biochemical parameters were monitored before and after the treatment. HFD rats developed obese insulin resistance which led to impaired dynamics balance and function of mitochondria, increased cardiac cell apoptosis and dysfunction. Although Mdivi1, M1 and combined treatment exerted similar cardiometabolic benefits in HFD rats, the combined therapy showed a greater reduction in mitochondrial reactive oxygen species (ROS). Mitochondrial fission inhibitor and fusion promoter exerted similar levels of cardioprotection in a pre-diabetic condition.

scholarly journals Actin-Related Protein 4 Interacts with PIE1 and Regulates Gene Expression in Arabidopsis

Genes ◽  
2021 ◽  
Vol 12 (4) ◽  
pp. 520
Author(s):  
Wenfeng Nie ◽  
Jinyu Wang
Keyword(s):  
Gene Expression ◽  
Plant Growth ◽  
Chromatin Remodeling ◽  
Leaf Size ◽  
Early Flowering ◽  
Physical Interaction ◽  
Loss Of Function ◽  
Single Nucleotide Change ◽  
Chromatin Remodeling Complex ◽  
Related Proteins

As essential structural components of ATP-dependent chromatin-remodeling complex, the nucleolus-localized actin-related proteins (ARPs) play critical roles in many biological processes. Among them, ARP4 is identified as an integral subunit of chromatin remodeling complex SWR1, which is conserved in yeast, humans and plants. It was shown that RNAi mediated knock-down of Arabidopsis thaliana ARP4 (AtARP4) could affect plant development, specifically, leading to early flowering. However, so far, little is known about how ARP4 functions in the SWR1 complex in plant. Here, we identified a loss-of-function mutant of AtARP4 with a single nucleotide change from glycine to arginine, which had significantly smaller leaf size. The results from the split luciferase complementation imaging (LCI) and yeast two hybrid (Y2H) assays confirmed its physical interaction with the scaffold and catalytic subunit of SWR1 complex, photoperiod-independent early flowering 1 (PIE1). Furthermore, mutation of AtARP4 caused altered transcription response of hundreds of genes, in which the number of up-regulated differentially expressed genes (DEGs) was much larger than those down-regulated. Although most DEGs in atarp4 are related to plant defense and response to hormones such as salicylic acid, overall, it has less overlapping with other swr1 mutants and the hta9 hta11 double-mutant. In conclusion, our results reveal that AtARP4 is important for plant growth and such an effect is likely attributed to its repression on gene expression, typically at defense-related loci, thus providing some evidence for the coordination of plant growth and defense, while the regulatory patterns and mechanisms are distinctive from other SWR1 complex components.

274 Tumoral and peripheral immunophenotype of refractory vs relapse to PD-(L)1 blockade in patients with advanced non-small cell lung cancer

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A299-A299
Author(s):  
Maria Ascierto ◽  
Matthew Hellmann ◽  
Nathan Standifer ◽  
Song Wu ◽  
Han Si ◽  
...  
Keyword(s):  
Gene Expression ◽  
Informed Consent ◽  
Checkpoint Inhibitors ◽  
Patient Response ◽  
Calcium Dependent ◽  
Immune Exhaustion ◽  
Previously Treated ◽  
Expression Pathways ◽  
Relapsed Disease ◽  
Written Informed Consent

BackgroundDespite the encouraging successes of immune checkpoint inhibitors, many patients do not benefit and are either refractory or relapse. The mechanisms of refractory or relapsed disease following PD-(L)1 blockade are largely unknown. To identify characteristics associated with refractory or relapsed disease we explored the immune and genomic landscape of samples derived from NSCLC patients who previously received PD-(L)1 blockade and had blood and fresh tumor biopsies collected at the time of progression.MethodsPatient response categories were defined prospectively; ‘refractory’ defined as progression within 16 weeks of initiating PD-(L)1 and ‘relapse’ defined as initial clinical benefit (CR, PR, SD) followed by progression. RNAseq (n=52) and PD-L1 IHC (n=22) were performed on tumor tissue. Immune profiling of whole blood was assessed using flow cytometry or Biomark HD (Fluidigm) gene expression panel (n=54 and n=62, respectively). Differential gene expression was defined as unadjusted p<0.05 and fold-difference >1.5. Pathways analysis was conducted by David tool. Patient samples were collected during screening for clinical trial of second line immunotherapy. Written informed consent was obtained from the patients for publication of this abstract.ResultsIn patients with NSCLC previously treated with PD-(L)1 blockade, tumors of relapsed patients were characterized by increased expression of genes associated with interferon signaling (e.g. CXCL9, SPIC, IFNg), immune suppression (e.g. ARG1, TGFB), immune exhaustion (e.g. ADORA2A), and increased PD-L1 expression (by gene expression and IHC). Refractory disease was associated with increased cadherin signaling and calcium-dependent-cell-adhesion gene expression pathways. In the periphery, reduced quantities of B cells and activated (HLA-DR+ or CD38+) or proliferating (Ki67+) CD8+ T cells were observed in refractory patients.ConclusionsThe tumor and peripheral compartments of patients with NSCLC previously treated with PD-(L)1 blockade differ based on prior response. Relapsed patients tend to have signals of sturdy immune activation and chronic inflammation thus ultimately leading to immune exhaustion. These results may help inform rational therapeutic strategies to overcome resistance to PD-(L)1 blockade in NSCLC.Trial RegistrationNCT02000947Ethics ApprovalResearch on human samples here analyzed have been performed in accordance with the Declaration of Helsinki.ConsentWritten informed consent was obtained from the patient for publication of this abstract.

scholarly journals Mitochondrial fission and mitophagy are independent mechanisms regulating ischemia/reperfusion injury in primary neurons

2021 ◽  
Vol 12 (5) ◽  
Author(s):  
Anthony R. Anzell ◽  
Garrett M. Fogo ◽  
Zoya Gurm ◽  
Sarita Raghunayakula ◽  
Joseph M. Wider ◽  
...  
Keyword(s):  
Cortical Neurons ◽  
Ischemia Reperfusion Injury ◽  
Mitochondrial Dynamics ◽  
Ischemia Reperfusion ◽  
Mitochondrial Fission ◽  
Conditional Knockout ◽  
Mitochondrial Morphology ◽  
Primary Neurons ◽  
Mitochondrial Fragmentation

AbstractMitochondrial dynamics and mitophagy are constitutive and complex systems that ensure a healthy mitochondrial network through the segregation and subsequent degradation of damaged mitochondria. Disruption of these systems can lead to mitochondrial dysfunction and has been established as a central mechanism of ischemia/reperfusion (I/R) injury. Emerging evidence suggests that mitochondrial dynamics and mitophagy are integrated systems; however, the role of this relationship in the context of I/R injury remains unclear. To investigate this concept, we utilized primary cortical neurons isolated from the novel dual-reporter mitochondrial quality control knockin mice (C57BL/6-Gt(ROSA)26Sortm1(CAG-mCherry/GFP)Ganl/J) with conditional knockout (KO) of Drp1 to investigate changes in mitochondrial dynamics and mitophagic flux during in vitro I/R injury. Mitochondrial dynamics was quantitatively measured in an unbiased manner using a machine learning mitochondrial morphology classification system, which consisted of four different classifications: network, unbranched, swollen, and punctate. Evaluation of mitochondrial morphology and mitophagic flux in primary neurons exposed to oxygen-glucose deprivation (OGD) and reoxygenation (OGD/R) revealed extensive mitochondrial fragmentation and swelling, together with a significant upregulation in mitophagic flux. Furthermore, the primary morphology of mitochondria undergoing mitophagy was classified as punctate. Colocalization using immunofluorescence as well as western blot analysis revealed that the PINK1/Parkin pathway of mitophagy was activated following OGD/R. Conditional KO of Drp1 prevented mitochondrial fragmentation and swelling following OGD/R but did not alter mitophagic flux. These data provide novel evidence that Drp1 plays a causal role in the progression of I/R injury, but mitophagy does not require Drp1-mediated mitochondrial fission.

scholarly journals When the Balance Tips: Dysregulation of Mitochondrial Dynamics as a Culprit in Disease

2021 ◽  
Vol 22 (9) ◽  
pp. 4617
Author(s):  
Styliana Kyriakoudi ◽  
Anthi Drousiotou ◽  
Petros P. Petrou
Keyword(s):  
Insulin Resistance ◽  
Mitochondrial Function ◽  
Human Disease ◽  
Molecular Mechanisms ◽  
Mitochondrial Dynamics ◽  
Mitochondrial Fusion ◽  
Mitochondrial Morphology ◽  
Disease Development ◽  
Pathological Conditions ◽  
Wide Range

Mitochondria are dynamic organelles, the morphology of which is tightly linked to their functions. The interplay between the coordinated events of fusion and fission that are collectively described as mitochondrial dynamics regulates mitochondrial morphology and adjusts mitochondrial function. Over the last few years, accruing evidence established a connection between dysregulated mitochondrial dynamics and disease development and progression. Defects in key components of the machinery mediating mitochondrial fusion and fission have been linked to a wide range of pathological conditions, such as insulin resistance and obesity, neurodegenerative diseases and cancer. Here, we provide an update on the molecular mechanisms promoting mitochondrial fusion and fission in mammals and discuss the emerging association of disturbed mitochondrial dynamics with human disease.

scholarly journals Syntaxin-17-Dependent Mitochondrial Dynamics is Essential for Protection Against Oxidative-Stress-Induced Apoptosis

Antioxidants ◽  
2019 ◽  
Vol 8 (11) ◽  
pp. 522 ◽  
Author(s):  
Wang ◽  
Xiao ◽  
Huang ◽  
Liu
Keyword(s):  
Oxidative Stress ◽  
Mitochondrial Dynamics ◽  
Mitochondrial Fission ◽  
Autophagic Flux ◽  
Wild Type ◽  
Dual Roles ◽  
Defective Mutant ◽  
U2os Cells ◽  
Induced Apoptosis

In this study, cell death induced by the oxidant tert-butylhydroperoxide (tBH) was observed in U2OS cells; this phenotype was rescued by Syntaxin 17 (STX17) knockout (KO) but the mechanism is unknown. STX17 plays dual roles in autophagosome–lysosome fusion and mitochondrial fission. However, the contribution of the two functions of STX17 to apoptosis has not been extensively studied. Here, we sought to dissect the dual roles of STX17 in oxidative-stress-induced apoptosis by taking advantage of STX17 knockout cells and an autophagosome–lysosome fusion defective mutant of STX17. We generated STX17 knockout U2OS cells using the clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) system and the STX17 knockout cells were reconstituted with wild-type STX17 and its autophagosome–lysosome fusion defective mutant. Autophagy was assessed by autophagic flux assay, Monomer red fluorescent protein (mRFP)–GFP–LC3 assay and protease protection assay. Golgi, endoplasmic reticulum (ER)/ER–Golgi intermediate compartment (ERGIC) and mitochondrial dynamics were examined by staining the different indicator proteins. Apoptosis was evaluated by caspase cleavage assay. The general reactive oxygen species (ROS) were detected by flow cytometry. In STX17 complete knockout cells, sealed autophagosomes were efficiently formed but their fusion with lysosomes was less defective. The fusion defect was rescued by wild-type STX17 but not the autophagosome–lysosome fusion defective mutant. No obvious defects in Golgi, ERGIC or ER dynamics were observed. Mitochondria were significantly elongated, supporting a role of STX17 in mitochondria fission and the elongation caused by STX17 KO was reversed by the autophagosome–lysosome fusion defective mutant. The clearance of protein aggregation was compromised, correlating with the autophagy defect but not with mitochondrial dynamics. This study revealed a mixed role of STX17 in autophagy, mitochondrial dynamics and oxidative stress response. STX17 knockout cells were highly resistant to oxidative stress, largely due to the function of STX17 in mitochondrial fission rather than autophagy.

scholarly journals Non-alcoholic fatty liver disease in mice with hepatocyte-specific deletion of mitochondrial fission factor

Diabetologia ◽  
2021 ◽  
Author(s):  
Yukina Takeichi ◽  
Takashi Miyazawa ◽  
Shohei Sakamoto ◽  
Yuki Hanada ◽  
Lixiang Wang ◽  
...  
Keyword(s):  
Er Stress ◽  
Mitochondrial Dynamics ◽  
Mitochondrial Fission ◽  
Primary Hepatocytes ◽  
Alcoholic Fatty Liver ◽  
Expression Of Genes ◽  
Specific Deletion

Abstract Aims/hypothesis Mitochondria are highly dynamic organelles continuously undergoing fission and fusion, referred to as mitochondrial dynamics, to adapt to nutritional demands. Evidence suggests that impaired mitochondrial dynamics leads to metabolic abnormalities such as non-alcoholic steatohepatitis (NASH) phenotypes. However, how mitochondrial dynamics are involved in the development of NASH is poorly understood. This study aimed to elucidate the role of mitochondrial fission factor (MFF) in the development of NASH. Methods We created mice with hepatocyte-specific deletion of MFF (MffLiKO). MffLiKO mice fed normal chow diet (NCD) or high-fat diet (HFD) were evaluated for metabolic variables and their livers were examined by histological analysis. To elucidate the mechanism of development of NASH, we examined the expression of genes related to endoplasmic reticulum (ER) stress and lipid metabolism, and the secretion of triacylglycerol (TG) using the liver and primary hepatocytes isolated from MffLiKO and control mice. Results MffLiKO mice showed aberrant mitochondrial morphologies with no obvious NASH phenotypes during NCD, while they developed full-blown NASH phenotypes in response to HFD. Expression of genes related to ER stress was markedly upregulated in the liver from MffLiKO mice. In addition, expression of genes related to hepatic TG secretion was downregulated, with reduced hepatic TG secretion in MffLiKO mice in vivo and in primary cultures of MFF-deficient hepatocytes in vitro. Furthermore, thapsigargin-induced ER stress suppressed TG secretion in primary hepatocytes isolated from control mice. Conclusions/interpretation We demonstrated that ablation of MFF in liver provoked ER stress and reduced hepatic TG secretion in vivo and in vitro. Moreover, MffLiKO mice were more susceptible to HFD-induced NASH phenotype than control mice, partly because of ER stress-induced apoptosis of hepatocytes and suppression of TG secretion from hepatocytes. This study provides evidence for the role of mitochondrial fission in the development of NASH. Graphical abstract

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