bims-mikwok 2022-06-26 papers

bims-mikwok

Biomed News

on Mitochondrial quality control

Issue of 2022‒06‒26
twenty papers selected by
Avinash N. Mukkala
University of Toronto

Mitochondria transplantation/transfer between single cells.
Inhibition of mitoNEET induces Pink1-Parkin-mediated mitophagy.
Mitochondrial metabolism and bioenergetic function in an anoxic isolated adult mouse cardiomyocyte model of in vivo cardiac ischemia-reperfusion injury.
Mitochondrial Quality Control in the Heart: The Balance between Physiological and Pathological Stress.
Mitochondrial-derived vesicles: Gatekeepers of mitochondrial response to oxidative stress.
Transcription factor nuclear factor erythroid 2 p45-related factor 2 (NRF2) ameliorates sepsis-associated acute kidney injury by maintaining mitochondrial homeostasis and improving the mitochondrial function.
Protocol for engineering and validating a synthetic mitochondrial intermembrane bridge in mammalian cells.
ULK1 mediated mitophagy prevents pathological cardiac remodelling and heart failure.
Assessment of Open Probability of the Mitochondrial Permeability Transition Pore in the Setting of Coenzyme Q Excess.
Tollip negatively regulates mitophagy by promoting the mitochondrial processing and cytoplasmic release of PINK1.
Effects of sugars, fatty acids and amino acids on cytosolic and mitochondrial hydrogen peroxide release from liver cells.
RIP3 Translocation into Mitochondria Promotes Mitofilin Degradation to Increase Inflammation and Kidney Injury after Renal Ischemia-Reperfusion.
Substrate binding in the mitochondrial ADP/ATP carrier is a step-wise process guiding the structural changes in the transport cycle.
TFAM downregulation promotes autophagy and ESCC survival through mtDNA stress-mediated STING pathway.
STING signaling sensing of DRP1-dependent mtDNA release in kupffer cells contributes to lipopolysaccharide-induced liver injury in mice.
Tankyrases inhibit innate antiviral response by PARylating VISA/MAVS and priming it for RNF146-mediated ubiquitination and degradation.
The role of ubiquitin signaling pathway on liver regeneration in rats.
Monitoring DNA polymerase β mitochondrial localization and dynamics.
Mitochondrial calcium uniporter promotes phagocytosis-dependent activation of the NLRP3 inflammasome.
Remote ischemic preconditioning causes transient cell-cycle arrest and renal protection by a NF-kB-dependent Sema5B pathway.

J Cereb Blood Flow Metab. 2022 Jun 21. 271678X221109685

Mitochondria transplantation/transfer between single cells.

Qin Hu, Jianfei Lu, Xiaohua Zhang, Ran Liu, Shao-Hua Yang.

  Mitochondrial transplantation/transfer has been increasingly recognized as a potential way for cell and tissue revitalization. In a recent study, Gabelein et al. reported a novel method for single cells mitochondria transplantation using "nanosyringe". This technique combines atomic force microscopy, optical microscopy, and nanofluidics that enable intra- and intercellular organelle micromanipulation and cell-to-cell mitochondria transplantation with up to 95% success rate. The transferred mitochondria fuse to the host mitochondrial network and donor mtDNA incorporate into the recipient mitochondrial genome. The nanosyringe technique provides a novel tool for future mitochondrial research to offer insight into mitochondrial replacement therapy for stroke and fundamental mitochondrial biology.

Keywords:  Mitochondria; mitochondrial transfer; mitochondrial transplantation; nanosyringe; revitalization

DOI:  https://doi.org/10.1177/0271678X221109685
BMB Rep. 2022 Jun 21. pii: 5590. [Epub ahead of print]

Inhibition of mitoNEET induces Pink1-Parkin-mediated mitophagy.

Seunghee Lee, Sangguk Lee, Seon-Jin Lee, Su Wol Chung.

MitoNEET, a mitochondrial outer membrane protein containing the Asn-Glu-Glu-Thr (NEET) sequence, controls the formation of intermitochondrial junctions and confers autophagy resistance. Moreover, mitoNEET as a mitochondrial substrate undergoes ubiquitination by activated Parkin during the initiation of mitophagy. Therefore, mitoNEET is linked to the regulation of autophagy and mitophagy. Mitophagy is the selective removal of the damaged or unnecessary mitochondria, which is crucial to sustaining mitochondrial quality control. In numerous human diseases, the accumulation of damaged mitochondria by impaired mitophagy has been observed. However, the therapeutic strategy targeting of mitoNEET as a mitophagy-enhancing mediator requires further research. Herein, we confirmed that mitophagy is indeed activated by mitoNEET inhibition. CCCP (carbonyl cyanide m-chlorophenyl hydrazone), which leads to mitochondrial depolarization, induces mitochondrial dysfunction and superoxide production. This, in turn, contributes to the induction of mitophagy; mitoNEET protein levels were initially increased before an increase in LC3-Ⅱ protein following CCCP treatment. Pharmacological inhibition of mitoNEET using mitoNEET Ligand-1 (NL-1) promoted accumulation of Pink1 and Parkin, which are mitophagy-associated proteins, and activation of mitochondria-lysosome crosstalk, in comparison to CCCP alone. Inhibition of mitoNEET using NL-1, or mitoNEET shRNA transfected into RAW264.7 cells, abrogated CCCP-induced ROS and mitochondrial cell death; additionally, it activated the expression of PGC-1α and SOD2, regulators of oxidative metabolism. In particular, the increase in PGC-1α, which is a major regulator of mitochondrial biogenesis, promotes mitochondrial quality control. These results indicated that mitoNEET is a potential therapeutic target in numerous human diseases to enhance mitophagy and protect cells by maintaining a network of healthy mitochondria.
Redox Biol. 2022 Jun 17. pii: S2213-2317(22)00140-9. [Epub ahead of print]54 102368

Mitochondrial metabolism and bioenergetic function in an anoxic isolated adult mouse cardiomyocyte model of in vivo cardiac ischemia-reperfusion injury.

Anja V Gruszczyk, Alva M Casey, Andrew M James, Hiran A Prag, Nils Burger, Georgina R Bates, Andrew R Hall, Fay M Allen, Thomas Krieg, Kourosh Saeb-Parsy, Michael P Murphy.

  Cell models of cardiac ischemia-reperfusion (IR) injury are essential to facilitate understanding, but current monolayer cell models poorly replicate the in vivo IR injury that occurs within a three-dimensional tissue. Here we show that this is for two reasons: the residual oxygen present in many cellular hypoxia models sustains mitochondrial oxidative phosphorylation; and the loss of lactate from cells into the incubation medium during ischemia enables cells to sustain glycolysis. To overcome these limitations, we incubated isolated adult mouse cardiomyocytes anoxically while inhibiting lactate efflux. These interventions recapitulated key markers of in vivo ischemia, notably the accumulation of succinate and the loss of adenine nucleotides. Upon reoxygenation after anoxia the succinate that had accumulated during anoxia was rapidly oxidized in association with extensive mitochondrial superoxide/hydrogen peroxide production and cell injury, mimicking reperfusion injury. This cell model will enable key aspects of cardiac IR injury to be assessed in vitro.

Keywords:  Cardiomyocytes; Hydrogen peroxide; Ischemia-reperfusion injury; Metabolism; Mitochondria

DOI:  https://doi.org/10.1016/j.redox.2022.102368
Biomedicines. 2022 Jun 10. pii: 1375. [Epub ahead of print]10(6):

Mitochondrial Quality Control in the Heart: The Balance between Physiological and Pathological Stress.

Giovanni Fajardo, Michael Coronado, Melia Matthews, Daniel Bernstein.

  Alterations in mitochondrial function and morphology are critical adaptations to cardiovascular stress, working in concert in an attempt to restore organelle-level and cellular-level homeostasis. Processes that alter mitochondrial morphology include fission, fusion, mitophagy, and biogenesis, and these interact to maintain mitochondrial quality control. Not all cardiovascular stress is pathologic (e.g., ischemia, pressure overload, cardiotoxins), despite a wealth of studies to this effect. Physiological stress, such as that induced by aerobic exercise, can induce morphologic adaptations that share many common pathways with pathological stress, but in this case result in improved mitochondrial health. Developing a better understanding of the mechanisms underlying alterations in mitochondrial quality control under diverse cardiovascular stressors will aid in the development of pharmacologic interventions aimed at restoring cellular homeostasis.

Keywords:  biogenesis; fission; fusion; mitochondria; mitophagy

DOI:  https://doi.org/10.3390/biomedicines10061375
Free Radic Biol Med. 2022 Jun 21. pii: S0891-5849(22)00460-9. [Epub ahead of print]

Mitochondrial-derived vesicles: Gatekeepers of mitochondrial response to oxidative stress.

Tingting Peng, Yinyin Xie, Hanqing Sheng, Cui Wang, Yajun Lian, Nanchang Xie.

  Mitochondrial quality control (MQC) mechanisms are a series of adaptive responses that ensure the relative stability of mitochondrial morphology, quantity, and quality to preserve cellular survival and function. While MQC mechanisms range from mitochondrial biogenesis and fusion/fission to mitophagy, mitochondrial-derived vesicles (MDVs) may represent an essential component of MQC. MDVs precede mitochondrial autophagy and serve as the first line of defense against oxidative stress by selectively transferring damaged mitochondrial substances to the lysosome for degradation. In fact, the function of MDVs is dependent on the cargo, the shuttle route, and the ultimate destination. Abnormal MDVs disrupt metabolite clearance and the immune response, predisposing to pathological conditions, including neurodegeneration, cardiovascular diseases, and cancers. Therefore, MDV regulation may be a potential therapeutic for the therapy of these diseases. In this review, we highlight recent advances in the study of MDVs and their misregulation in various diseases from the perspectives of formation, cargo selection, regulation, and transportation.

Keywords:  Aging; Diseases; Mitochondria; Mitochondrial quality control; Mitochondrial-derived vesicle; Therapeutics

DOI:  https://doi.org/10.1016/j.freeradbiomed.2022.06.233
Eur J Histochem. 2022 Jun 21. 66(3):

Transcription factor nuclear factor erythroid 2 p45-related factor 2 (NRF2) ameliorates sepsis-associated acute kidney injury by maintaining mitochondrial homeostasis and improving the mitochondrial function.

Zhijiang Chen, Huili Wang, Bin Hu, Xinxin Chen, Meiyu Zheng, Lili Liang, Juanjuan Lyu, Qiyi Zeng.

Mitochondrial dysfunction has a role in sepsis-associated acute kidney injury (S-AKI), so the restoration of normal mitochondrial homeostasis may be an effective treatment strategy. Transcription factor nuclear factor erythroid 2 p45-related factor 2 (NRF2) is a main regulator of cell-redox homeostasis, and recent studies reported that NRF2 activation helped to preserve mitochondrial morphology and function under conditions of stress. However, the role of NRF2 in the process of S-AKI is still not well understood. The present study investigated whether NRF2 regulates mitochondrial homeostasis and influences mitochondrial function in S-AKI. We demonstrated activation of NRF2 in an in vitro model: lipopolysaccharide (LPS) challenge of ductal epithelial cells of rat renal tubules (NRK-52e cells), and an in vivo model: cecal ligation and puncture (CLP) of rats. Over-expression of NRF2 attenuated oxidative stress, apoptosis, and the inflammatory response; enhanced mitophagy and mitochondrial biogenesis; and mitigated mitochondrial damage in the in vitro model. In vivo experiments showed that rats treated with an NRF2 agonist had higher adenosine triphosphate (ATP) levels, lower blood urea nitrogen and creatinine levels, fewer renal histopathological changes, and higher expression of mitophagy-related proteins [PTEN-induced putative kinase 1 (PINK1), parkin RBR E3 ubiquitin protein ligase (PRKN), microtubule-associated protein 1 light chain 3 II (LC3 II)] and mitochondrial biogenesis-related proteins [peroxisome proliferator-activated receptor γ coactivator-1 (PGC-1α) and mitochondrial transcription factor A (TFAM)]. Electron microscopy of kidney tissues showed that mitochondrial damage was alleviated by treatment with an NRF2 agonist, and the opposite response occurred upon treatment with an NRF2 antagonist. Overall, our findings suggest that mitochondria have an important role in the pathogenesis of S-AKI, and that NRF2 activation restored mitochondrial homeostasis and function in the presence of this disease. This mitochondrial pathway has the potential to be a novel therapeutic target for the treatment of S-AKI.

DOI: https://doi.org/10.4081/ejh.2022.3412
STAR Protoc. 2022 Sep 16. 3(3): 101454

Protocol for engineering and validating a synthetic mitochondrial intermembrane bridge in mammalian cells.

Gunjan Purohit, Martonio Ponte Viana, Oleh Khalimonchuk.

  Membrane contact sites are recognized as critical means of intercompartmental communication. Here, we describe a protocol for engineering and validating a synthetic bridge between the inner and outer mitochondrial membranes to support functioning of the endogenous mitochondrial contact site and cristae organizing system (MICOS). A chimeric protein, MitoT, is stably expressed in cultured mammalian cells to bridge the mitochondrial membranes. This approach can be a valuable tool to study the function of the MICOS complex and associated proteins. For complete details on the use and execution of this protocol, please refer to Viana et al. (2021).

Keywords:  Biotechnology and bioengineering; Cell Biology; Cell Membrane; Cell culture; Cell isolation; Flow Cytometry/Mass Cytometry; Metabolism; Microscopy; Molecular Biology

DOI:  https://doi.org/10.1016/j.xpro.2022.101454
Cardiovasc Res. 2022 Jun 21. pii: cvac101. [Epub ahead of print]

ULK1 mediated mitophagy prevents pathological cardiac remodelling and heart failure.

Rimpy Dhingra, Lorrie A Kirshenbaum.

  The defects in mitochondrial clearance mechanisms can trigger adverse cardiac remodeling and severely impair cardiac performance. A new study identifies Ulk1/Rab9 mediated alternative mitophagy to be important for mitochondrial clearance in heart under pressure overload conditions. Moreover, the defects in ULK1 mediated alternative mitophagy resulted in accumulation of damaged mitochondria, severe hypertrophy, fibrosis, and cardiac dysfunction in response to TAC induced pressure overload. The findings highlight Ulk1/Rab9 mediated alternative mitophagy as a prominent mode of mitophagy and quality control in response to pressure overload hypertrophy.

Keywords:  Heart failure; Hypertrophy; Mitochondria; Mitophagy; Pressure-overload

DOI:  https://doi.org/10.1093/cvr/cvac101
J Vis Exp. 2022 Jun 01.

Assessment of Open Probability of the Mitochondrial Permeability Transition Pore in the Setting of Coenzyme Q Excess.

Keren K Griffiths, Aili Wang, Richard J Levy.

The mitochondrial permeability transition pore (mPTP) is a voltage-gated, nonselective, inner mitochondrial membrane (IMM) mega-channel important in health and disease. The mPTP mediates leakage of protons across the IMM during low-conductance opening and is specifically inhibited by cyclosporine A (CsA). Coenzyme Q (CoQ) is a regulator of the mPTP, and tissue-specific differences have been found in CoQ content and open probability of the mPTP in forebrain and heart mitochondria in a newborn mouse model of fragile X syndrome (FXS, Fmr1 knockout). We developed a technique to determine the voltage threshold for mPTP opening in this mutant strain, exploiting the role of the mPTP as a proton leak channel. To do so, oxygen consumption and membrane potential (ΔΨ) were simultaneously measured in isolated mitochondria using polarography and a tetraphenylphosphonium (TPP+) ion-selective electrode during leak respiration. The threshold for mPTP opening was determined by the onset of CsA-mediated inhibition of proton leak at specific membrane potentials. Using this approach, differences in voltage gating of the mPTP were precisely defined in the context of CoQ excess. This novel technique will permit future investigation for enhancing the understanding of physiological and pathological regulation of low-conductance opening of the mPTP.

DOI: https://doi.org/10.3791/63646
BMB Rep. 2022 Jun 21. pii: 5642. [Epub ahead of print]

Tollip negatively regulates mitophagy by promoting the mitochondrial processing and cytoplasmic release of PINK1.

Woo Hyun Shin, Kwang Chul Chung.

PTEN-induced putative kinase 1 (PINK1) is a serine/threonine kinase that phosphorylates several substrates and exerts neuroprotective effects against stress-induced apoptotic cell death. Mutations in PINK1 have been linked to autosomal recessive forms of Parkinson's disease (PD). Mitophagy is a type of autophagy that selectively promotes mitochondrial turnover and prevents the accumulation of dysfunctional mitochondria to maintain cellular homeostasis. Toll-interacting protein (Tollip) was initially identified as a negative regulator of IL-1β receptor signaling, suppressing inflammatory TLR signaling cascades. Recently, Tollip has been reported to play a role in autophagy and is implicated in neurodegeneration. In this study, we determined whether Tollip was functionally linked to PINK1-mediated mitophagy. Our results demonstrated that Tollip promoted the mitochondrial processing of PINK1 and altered the localization of PINK1, predominantly to the cytosol. This action was attributed to increased binding of PINK1 to mitochondrial processing peptidase β (MPPβ) and the subsequent increase in MPPβ-mediated mitochondrial PINK1 cleavage. Furthermore, Tollip suppressed mitophagy following carbonyl cyanide m-chlorophenylhydrazone-induced mitochondrial dysfunction. These findings suggest that Tollip inhibits mitophagy via the PINK1/parkin pathway upon mitochondrial damage, leading to the blockade of PINK1-mediated neuroprotection.
Free Radic Biol Med. 2022 Jun 16. pii: S0891-5849(22)00452-X. [Epub ahead of print]188 92-102

Effects of sugars, fatty acids and amino acids on cytosolic and mitochondrial hydrogen peroxide release from liver cells.

Jingqi Fang, Yini Zhang, Akos A Gerencser, Martin D Brand.

  The rates of formation of superoxide and hydrogen peroxide at different electron-donating sites in isolated mitochondria are critically dependent on the substrates that are added, through their effects on the reduction level of each site and the components of the protonmotive force. However, in intact cells the acute effects of added substrates on different sites of cytosolic and mitochondrial hydrogen peroxide production are unclear. Here we tested the effects of substrate addition on cytosolic and mitochondrial hydrogen peroxide release from intact AML12 liver cells. In 30-min starved cells replete with endogenous substrates, addition of glucose, fructose, palmitate, alanine, leucine or glutamine had no effect on the rate or origin of cellular hydrogen peroxide release. However, following 150-min starvation of the cells to deplete endogenous glycogen (and other substrates), cellular hydrogen peroxide production, particularly from NADPH oxidases (NOXs), was decreased, GSH/GSSH ratio increased, and antioxidant gene expression was unchanged. Addition of glucose or glutamine (but not the other substrates) increased hydrogen peroxide release. There were similar relative increases from each of the three major sites of production: mitochondrial sites IQ and IIIQo, and cytosolic NOXs. Glucose supplementation also restored ATP production and mitochondrial NAD reduction level, suggesting that the increased rates of hydrogen peroxide release from the mitochondrial sites were driven by increases in the protonmotive force and the degree of reduction of the electron transport chain. Long-term (24 h) glucose or glutamine deprivation also diminished hydrogen peroxide release rate, ATP production rate and (for glucose deprivation) NAD reduction level. We conclude that the rates of superoxide and hydrogen peroxide production from mitochondrial sites in liver cells are insensitive to extra added substrates when endogenous substrates are not depleted, but these rates are decreased when endogenous substrates are lowered by 150 min of starvation, and can be enhanced by restoring glucose or glutamine supply through improvements in mitochondrial energetic state.

Keywords:  Liver; Mitochondria; NOX; ROS; Site III(Qo); Site IQ

DOI:  https://doi.org/10.1016/j.freeradbiomed.2022.06.225
Cells. 2022 Jun 11. pii: 1894. [Epub ahead of print]11(12):

RIP3 Translocation into Mitochondria Promotes Mitofilin Degradation to Increase Inflammation and Kidney Injury after Renal Ischemia-Reperfusion.

Yansheng Feng, Abdulhafiz Imam Aliagan, Nathalie Tombo, Derrick Draeger, Jean C Bopassa.

  The receptor-interacting protein kinase 3 (RIP3) has been reported to regulate programmed necrosis-necroptosis forms of cell death with important functions in inflammation. We investigated whether RIP3 translocates into mitochondria in response to renal ischemia-reperfusion (I/R) to interact with inner mitochondrial protein (Mitofilin) and promote mtDNA release into the cytosol. We found that release of mtDNA activates the cGAS-STING pathway, leading to increased nuclear transcription of pro-inflammatory markers that exacerbate renal I/R injury. Monolateral C57/6N and RIP3-/- mice kidneys were subjected to 60 min of ischemia followed by either 12, 24, or 48 h of reperfusion. In WT mice, we found that renal I/R injury increased RIP3 levels, as well as its translocation into mitochondria. We observed that RIP3 interacts with Mitofilin, likely promoting its degradation, resulting in increased mitochondria damage and mtDNA release, activation of the cGAS-STING-p65 pathway, and increased transcription of pro-inflammatory markers. All of these effects observed in WT mice were decreased in RIP3-/- mice. In HK-2, RIP3 overexpression or Mitofilin knockdown increased cell death by activating the cGAS-STING-p65 pathway. Together, this study point to an important role of the RIP3-Mitofilin axis in the initiation and development of renal I/R injury.

Keywords:  acute kidney injury (AKI); cGAS–STING–p65 pathway; inflammation; inner mitochondrial membrane protein (immt; mitochondrial dysfunction; mitochondrial structural integrity and function; mitofilin); mtDNA release; receptor-interacting protein kinase 3 (RIP3)

DOI:  https://doi.org/10.3390/cells11121894
Nat Commun. 2022 Jun 23. 13(1): 3585

Substrate binding in the mitochondrial ADP/ATP carrier is a step-wise process guiding the structural changes in the transport cycle.

Vasiliki Mavridou, Martin S King, Sotiria Tavoulari, Jonathan J Ruprecht, Shane M Palmer, Edmund R S Kunji.

Mitochondrial ADP/ATP carriers import ADP into the mitochondrial matrix and export ATP to the cytosol to fuel cellular processes. Structures of the inhibited cytoplasmic- and matrix-open states have confirmed an alternating access transport mechanism, but the molecular details of substrate binding remain unresolved. Here, we evaluate the role of the solvent-exposed residues of the translocation pathway in the process of substrate binding. We identify the main binding site, comprising three positively charged and a set of aliphatic and aromatic residues, which bind ADP and ATP in both states. Additionally, there are two pairs of asparagine/arginine residues on opposite sides of this site that are involved in substrate binding in a state-dependent manner. Thus, the substrates are directed through a series of binding poses, inducing the conformational changes of the carrier that lead to their translocation. The properties of this site explain the electrogenic and reversible nature of adenine nucleotide transport.

DOI: https://doi.org/10.1038/s41467-022-31366-5
Oncogene. 2022 Jun 24.

TFAM downregulation promotes autophagy and ESCC survival through mtDNA stress-mediated STING pathway.

Yujia Li, Qi Yang, Hui Chen, Xiaotian Yang, Jingru Han, Xiaojuan Yao, Xiajie Wei, Jiaoyang Si, Huanling Yao, Hongliang Liu, Lixin Wan, Hushan Yang, Yanming Wang, Dengke Bao.

The dynamics of mitochondrial biogenesis regulation is critical in maintaining cellular homeostasis for immune regulation and tumor prevention. Here, we report that mitochondrial biogenesis disruption through TFAM reduction significantly impairs mitochondrial function, induces autophagy, and promotes esophageal squamous cell carcinoma (ESCC) growth. We found that TFAM protein reduction promotes mitochondrial DNA (mtDNA) release into the cytosol, induces cytosolic mtDNA stress, subsequently activates the cGAS-STING signaling pathway, thereby stimulating autophagy and ESCC growth. STING depletion or mtDNA degradation by DNase I abrogates mtDNA stress response, attenuates autophagy, and decreases the growth of TFAM depleted cells. In addition, autophagy inhibitor also ameliorates mitochondrial dysfunction-induced activation of the cGAS-STING signaling pathway and ESCC growth. In conclusion, our results indicate that mtDNA stress induced by mitochondria biogenesis perturbation activates the cGAS-STING pathway and autophagy to promote ESCC growth, revealing an underappreciated therapeutic strategy for ESCC.

DOI: https://doi.org/10.1038/s41388-022-02365-z
Redox Biol. 2022 Jun 15. pii: S2213-2317(22)00139-2. [Epub ahead of print]54 102367

STING signaling sensing of DRP1-dependent mtDNA release in kupffer cells contributes to lipopolysaccharide-induced liver injury in mice.

Qin Zhang, Jiayi Wei, Zhuanhua Liu, Xiaoxia Huang, Maomao Sun, Wujiang Lai, Zhenfeng Chen, Jie Wu, Yanjia Chen, Xiaohua Guo, Qiaobing Huang.

  Aberrant pro-inflammatory activation of Kupffer cells (KCs) is strongly involved in the pathogenesis of septic liver injury. Recent evidence indicates the crucial roles of excessive stimulator of interferon genes (STING) signaling activation during sepsis. However, the role of STING signaling in septic liver injury remains unclear. In this study, we demonstrated that STING signaling was markedly activated in KCs isolated from wild type mice after lipopolysaccharide (LPS) treatment. STING deficiency effectively protected liver function, attenuated systemic inflammatory response and decreased mortality in LPS-treated mice, which were aggravated by STING agonist (DMXAA). Importantly, STING signaling activation in KCs contributed to LPS-induced liver injury through promoting hepatocyte death. Mechanistically, STING signaling could be activated by release of mitochondrial DNA (mtDNA) through dynamin-related protein 1 (DRP1)-dependent mitochondrial fission in LPS-treated KCs. Additionally, LPS stimulation enhanced DRP1-dependent mitochondrial ROS production, which promoted the leak of mtDNA into the cytosol and subsequent STING signaling activation in KCs. The in vivo experiments showed that pharmacological inhibition of DRP1 with Mdivi-1 partially prevented the activation of STING signaling in KCs isolated from LPS-challenged mice, as well as alleviated liver injury and inhibited systemic inflammatory response. In summary, our study comprehensively confirmed that STING signaling senses the DRP1-dependent release of mtDNA in KCs and its activation might play a key role in LPS-induced liver injury, which offers new sights and therapeutic targets for management of septic liver injury.

Keywords:  DRP1; Kupffer cell; LPS; Liver injury; STING; mtDNA

DOI:  https://doi.org/10.1016/j.redox.2022.102367
Proc Natl Acad Sci U S A. 2022 Jun 28. 119(26): e2122805119

Tankyrases inhibit innate antiviral response by PARylating VISA/MAVS and priming it for RNF146-mediated ubiquitination and degradation.

Yan-Ran Xu, Meng-Ling Shi, Yu Zhang, Na Kong, Cong Wang, Yi-Feng Xiao, Shi-Shen Du, Qi-Yun Zhu, Cao-Qi Lei.

  During viral infection, sensing of viral RNA by retinoic acid-inducible gene-I-like receptors (RLRs) initiates an antiviral innate immune response, which is mediated by the mitochondrial adaptor protein VISA (virus-induced signal adaptor; also known as mitochondrial antiviral signaling protein [MAVS]). VISA is regulated by various posttranslational modifications (PTMs), such as polyubiquitination, phosphorylation, O-linked β-d-N-acetylglucosaminylation (O-GlcNAcylation), and monomethylation. However, whether other forms of PTMs regulate VISA-mediated innate immune signaling remains elusive. Here, we report that Poly(ADP-ribosyl)ation (PARylation) is a PTM of VISA, which attenuates innate immune response to RNA viruses. Using a biochemical purification approach, we identified tankyrase 1 (TNKS1) as a VISA-associated protein. Viral infection led to the induction of TNKS1 and its homolog TNKS2, which translocated from cytosol to mitochondria and interacted with VISA. TNKS1 and TNKS2 catalyze the PARylation of VISA at Glu137 residue, thereby priming it for K48-linked polyubiquitination by the E3 ligase Ring figure protein 146 (RNF146) and subsequent degradation. Consistently, TNKS1, TNKS2, or RNF146 deficiency increased the RNA virus-triggered induction of downstream effector genes and impaired the replication of the virus. Moreover, TNKS1- or TNKS2-deficient mice produced higher levels of type I interferons (IFNs) and proinflammatory cytokines after virus infection and markedly reduced virus loads in the brains and lungs. Together, our findings uncover an essential role of PARylation of VISA in virus-triggered innate immune signaling, which represents a mechanism to avoid excessive harmful immune response.

Keywords:  PARylation; RNA virus; TNKS; VISA; innate immune response

DOI:  https://doi.org/10.1073/pnas.2122805119
Mol Cell Biochem. 2022 Jun 24.

The role of ubiquitin signaling pathway on liver regeneration in rats.

Ayse Ozmen Yaylaci, Mediha Canbek.

  The ubiquitin signalling pathway is a large system associated with numerous intracellular mechanisms. However, its function in the liver regeneration process remains unknown. This particular study investigates the intracellular effect mechanisms of the ubiquitin signalling pathway. It also determines the differences in the expression of 88 genes belonging to the ubiquitin pathway using the RT-PCR array method. To conduct this research, three genes-that differed in the expression analysis were selected. Moreover, their proteins were analysed by western blot analysis while using Ki67 immunohistochemical analysis that determines proliferation rates by hour. It was determined that BRCA1 and BARD1, which are effective in DNA repair, play an active role at PH24. Similarly, Ube2t expression, which belongs to the Fanconi anaemia pathway associated with DNA repair, was also found to be high at PH12-72 h. In addition, it was revealed that the expressions of Anapc2, Anapc11, Cdc20 belonging to the APC/CCdc20 complex, which participate in cell cycle regulation, differed at different hours after PH. Expression of Mul1, which is involved in mitochondrial biogenesis and mitophagy mechanisms, peaked at PH12 under the observation. Considering the Mul1 gene expression difference, MUL1-mediated mitophagy and mitochondrial fission mechanism may be associated with liver regeneration. It was also determined that PARKIN-mediated mitophagy mechanisms are not active in 0-72 h of liver regeneration since PARKIN expression did not show a significant change in PH groups.

Keywords:  APC/C; Fanconi anaemia pathway; Homologous recombination repair; Liver regeneration; Mitochondrial dynamics; Ubiquitin proteasome system

DOI:  https://doi.org/10.1007/s11010-022-04482-5
DNA Repair (Amst). 2022 Jun 11. pii: S1568-7864(22)00090-8. [Epub ahead of print]116 103357

Monitoring DNA polymerase β mitochondrial localization and dynamics.

Julie K Horton, Agnes K Janoshazi, Cristina A Nadalutti, Ming-Lang Zhao, Donna F Stefanick, Samuel H Wilson.

  Mouse fibroblasts lacking (null) DNA polymerase β (pol β) were transfected with fluorescently tagged pol β and stained with biomarkers to allow visualization within living cells by confocal microscopy. Transient transfection resulted in varying pol β expression levels. Separating cells into three groups based on pol β fluorescence intensity and morphological distribution, permitted analysis of the concentration dependence and spatial distribution of cytoplasmic pol β. Colocalization between pol β and mitochondria was pol β concentration dependent. A decrease in overlap with nucleoids containing mitochondrial DNA (mtDNA) was observed at the highest pol β intensity where pol β exhibits a tubular appearance, suggesting the ability to load elevated levels of pol β into mitochondria readily available for relocation to damaged mtDNA. The dynamics of pol β and mitochondrial nucleoids were followed by confocal recording of time series images. Two populations of mitochondrial nucleoids were observed, with and without pol β. Micro-irradiation, known to form DNA single-strand breaks, in a line across nucleus and cytoplasm of pol β stably transfected cells enhanced apparent localization of pol β with mitochondria in the perinuclear region of the cytoplasm near the nuclear membrane. Exposure of pol β expressing cells to H2O2 resulted in a time-dependent increase in cytoplasmic pol β observed by immunofluorescence analysis of fixed cells. Further screening revealed increased levels of colocalization of pol β with a mitochondrial probe and an increase in oxidative DNA damage in the cytoplasm. ELISA quantification confirmed an increase of an oxidative mitochondrial base lesion, 7,8-dihydro-8-oxoguanine, after H2O2 treatment. Taken together, the results suggest that pol β is recruited to mitochondria in response to oxidatively-induced mtDNA damage to participate in mtDNA repair.

Keywords:  DNA damage; DNA polymerase β; DNA repair; Laser micro-irradiation; Mitochondria; Mitochondrial DNA; Mitochondrial nucleoids

DOI:  https://doi.org/10.1016/j.dnarep.2022.103357
Proc Natl Acad Sci U S A. 2022 Jun 28. 119(26): e2123247119

Mitochondrial calcium uniporter promotes phagocytosis-dependent activation of the NLRP3 inflammasome.

Hong Dong, Bao Zhao, Jianwen Chen, Zihao Liu, Xinghui Li, Lupeng Li, Haitao Wen.

  Mitochondria, a highly metabolically active organelle, have been shown to play an essential role in regulating innate immune function. Mitochondrial Ca2+ uptake via the mitochondrial Ca2+ uniporter (MCU) is an essential process regulating mitochondrial metabolism by targeting key enzymes involved in the tricarboxylic acid cycle (TCA). Accumulative evidence suggests MCU-dependent mitochondrial Ca2+ signaling may bridge the metabolic reprogramming and regulation of immune cell function. However, the mechanism by which MCU regulates inflammation and its related disease remains elusive. Here we report a critical role of MCU in promoting phagocytosis-dependent activation of NLRP3 (nucleotide-binding domain, leucine-rich repeat containing family, pyrin domain-containing 3) inflammasome by inhibiting phagolysosomal membrane repair. Myeloid deletion of MCU (McuΔmye) resulted in an attenuated phagolysosomal rupture, leading to decreased caspase-1 cleavage and interleukin (IL)-1β release, in response to silica or alum challenge. In contrast, other inflammasome agonists such as adenosine triphosphate (ATP), nigericin, poly(dA:dT), and flagellin induced normal IL-1β release in McuΔmye macrophages. Mechanistically, we demonstrated that decreased NLRP3 inflammasome activation in McuΔmye macrophages was caused by improved phagolysosomal membrane repair mediated by ESCRT (endosomal sorting complex required for transport)-III complex. Furthermore, McuΔmye mice showed a pronounced decrease in immune cell recruitment and IL-1β production in alum-induced peritonitis, a typical IL-1-dependent inflammation model. In sum, our results identify a function of MCU in promoting phagocytosis-dependent NLRP3 inflammatory response via an ESCRT-mediated phagolysosomal membrane repair mechanism.

Keywords:  ESCRT; MCU; inflammasome; phagosome

DOI:  https://doi.org/10.1073/pnas.2123247119
JCI Insight. 2022 Jun 21. pii: e158523. [Epub ahead of print]

Remote ischemic preconditioning causes transient cell-cycle arrest and renal protection by a NF-kB-dependent Sema5B pathway.

Jan Rossaint, Melanie Meersch, Katharina Thomas, Sina Mersmann, Martin Lehmann, Jennifer Skupski, Tobias Tekath, Peter Rosenberger, John A Kellum, Hermann Pavenstädt, Alexander Zarbock.

  Acute kidney injury increases morbidity and mortality and previous studies have shown that remote ischemic preconditioning (RIPC) reduces the risk of acute kidney injury after cardiac surgery. RIPC increases urinary HMGB1 (high mobility group box protein-1) levels in patients which correlates with kidney protection. Here, we show that RIPC reduces renal ischemia-reperfusion injury and improves kidney function in mice. Mechanistically, RIPC increases HMGB1 levels in the plasma and urine and HMGB1 binds to Toll-like receptor 4 (TLR4) on renal tubular epithelial cells, inducing transcriptomic modulation of renal tubular epithelial cells and providing renal protection, whereas TLR4 activation on non-renal cells was shown to contribute to renal injury. This protection is mediated by activation of induction of AMPK⍺ and NF-kB, which induces the upregulation of Sema5b that triggers a transient, protective G1 cell-cycle arrest. In cardiac surgery patients at high risk for postoperative acute kidney injury, increased HMGB1 and Sema5b levels after RIPC were associated with renal protection after surgery. The results may help to develop future clinical treatment options for acute kidney injury.

Keywords:  Immunology; Nephrology; Neutrophils

DOI:  https://doi.org/10.1172/jci.insight.158523