bims-mitlys Biomed News
on Mitochondria and Lysosomes
Issue of 2021‒10‒24
fourteen papers selected by
Nicoletta Plotegher
University of Padova
  1. Precise control of mitophagy through ubiquitin proteasome system and deubiquitin proteases and their dysfunction in Parkinson's disease.
  2. The dynamics of mitochondrial autophagy at the initiation stage.
  3. GAK and PRKCD are positive regulators of PRKN-independent mitophagy.
  4. Mitophagy reporter mouse analysis reveals increased mitophagy activity in disuse-induced muscle atrophy.
  5. Quantifying inter-organelle membrane contact sites using proximity ligation assay in fixed optic nerve sections.
  6. Alleviation of CCCP injury by Augmenter of liver regeneration via the PINK1/Parkin pathway-dependent mitophagy.
  7. A three-organelle complex made by wrappER contacts with peroxisome and mitochondria responds to liver lipid flux changes.
  8. Exercise-Induced Benefits for Alzheimer's Disease by Stimulating Mitophagy and Improving Mitochondrial Function.
  9. Apelin-13-Mediated AMPK ameliorates endothelial barrier dysfunction in acute lung injury mice via improvement of mitochondrial function and autophagy.
  10. Mitochondrial Dysfunction, Protein Misfolding and Neuroinflammation in Parkinson's Disease: Roads to Biomarker Discovery.
  11. Full-coverage regulations of autophagy by ROS: from induction to maturation.
  12. Quantification of organelle contact sites by split-GFP-based contact site sensors (SPLICS) in living cells.
  13. New Insights into Molecular Mechanisms Mediating Adaptation to Exercise; A Review Focusing on Mitochondrial Biogenesis, Mitochondrial Function, Mitophagy and Autophagy.
  14. Lysosomal Zn2+ release triggers rapid, mitochondria-mediated, non-apoptotic cell death in metastatic melanoma.
  1. BMB Rep. 2021 Oct 22. pii: 5426. [Epub ahead of print]
    Precise control of mitophagy through ubiquitin proteasome system and deubiquitin proteases and their dysfunction in Parkinson's disease.
    Ga Hyun Park, Joon Hyung Park, Kwang Chul Chung.
      Parkinson's disease (PD) is one of the most common neurodegenerative diseases in the elderly population and is caused by the loss of dopaminergic neurons. PD has been predominantly attributed to mitochondrial dysfunction. The structural alteration of α-synuclein triggers toxic oligomer formation in the neurons, which greatly contributes to PD. In this article, we discuss the role of several familial PD-related proteins, such as α-synuclein, DJ-1, LRRK2, PINK1, and parkin in mitophagy, which entails a selective degradation of mitochondria via autophagy. Defective changes in mitochondrial dynamics and their biochemical and functional interaction induce the formation of toxic α-synuclein-containing protein aggregates in PD. In addition, these gene products play an essential role in ubiquitin proteasome system (UPS)-mediated proteolysis as well as mitophagy. Interestingly, a few deubiquitinating enzymes (DUBs) additionally modulate these two pathways negatively or positively. Based on these findings, we summarize the close relationship between several DUBs and the precise modulation of mitophagy. For example, the USP8, USP10, and USP15, among many DUBs are reported to specifically regulate the K48- or K63-linked de-ubiquitination reactions of several target proteins associated with the mitophagic process, in turn upregulating the mitophagy and protecting neuronal cells from α-synuclein-derived toxicity. In contrast, USP30 inhibits mitophagy by opposing parkin-mediated ubiquitination of target proteins. Furthermore, the association between these changes and PD pathogenesis will be discussed. Taken together, although the functional roles of several PD-related genes have yet to be fully understood, they are substantially associated with mitochondrial quality control as well as UPS. Therefore, a better understanding of their relationship provides valuable therapeutic clues for appropriate management strategies.
  2. Biochem Soc Trans. 2021 Oct 19. pii: BST20210272. [Epub ahead of print]
    The dynamics of mitochondrial autophagy at the initiation stage.
    Nicholas T Ktistakis.
      The pathway of mitochondrial-specific autophagy (mitophagy, defined here as the specific elimination of mitochondria following distinct mitochondrial injuries or developmental/metabolic alterations) is important in health and disease. This review will be focussed on the earliest steps of the pathway concerning the mechanisms and requirements for initiating autophagosome formation on a mitochondrial target. More specifically, and in view of the fact that we understand the basic mechanism of non-selective autophagy and are beginning to reshape this knowledge towards the pathways of selective autophagy, two aspects of mitophagy will be covered: (i) How does a machinery normally working in association with the endoplasmic reticulum (ER) to make an autophagosome can also do so at a site distinct from the ER such as on the surface of the targeted cargo? and (ii) how does the machinery deal with cargo of multiple sizes?
    Keywords:  autophagy; endoplasmic reticulum; imaging techniques; mitochondria
    DOI:  https://doi.org/10.1042/BST20210272
  3. Nat Commun. 2021 Oct 20. 12(1): 6101
    GAK and PRKCD are positive regulators of PRKN-independent mitophagy.
    Michael J Munson, Benan J Mathai, Matthew Yoke Wui Ng, Laura Trachsel-Moncho, Laura R de la Ballina, Sebastian W Schultz, Yahyah Aman, Alf H Lystad, Sakshi Singh, Sachin Singh, Jørgen Wesche, Evandro F Fang, Anne Simonsen.
      The mechanisms involved in programmed or damage-induced removal of mitochondria by mitophagy remains elusive. Here, we have screened for regulators of PRKN-independent mitophagy using an siRNA library targeting 197 proteins containing lipid interacting domains. We identify Cyclin G-associated kinase (GAK) and Protein Kinase C Delta (PRKCD) as regulators of PRKN-independent mitophagy, with both being dispensable for PRKN-dependent mitophagy and starvation-induced autophagy. We demonstrate that the kinase activity of both GAK and PRKCD are required for efficient mitophagy in vitro, that PRKCD is present on mitochondria, and that PRKCD facilitates recruitment of ULK1/ATG13 to early autophagic structures. Importantly, we demonstrate in vivo relevance for both kinases in the regulation of basal mitophagy. Knockdown of GAK homologue (gakh-1) in C. elegans or knockout of PRKCD homologues in zebrafish led to significant inhibition of basal mitophagy, highlighting the evolutionary relevance of these kinases in mitophagy regulation.
    DOI:  https://doi.org/10.1038/s41467-021-26331-7
  4. J Cell Physiol. 2021 Nov;236(11): 7612-7624
    Mitophagy reporter mouse analysis reveals increased mitophagy activity in disuse-induced muscle atrophy.
    Shun-Ichi Yamashita, Masanao Kyuuma, Keiichi Inoue, Yuki Hata, Ryu Kawada, Masaki Yamabi, Yasuyuki Fujii, Junko Sakagami, Tomoyuki Fukuda, Kentaro Furukawa, Satoshi Tsukamoto, Tomotake Kanki.
      Muscle disuse induces atrophy through increased reactive oxygen species (ROS) released from damaged mitochondria. Mitophagy, the autophagic degradation of mitochondria, is associated with increased ROS production. However, the mitophagy activity status during disuse-induced muscle atrophy has been a subject of debate. Here, we developed a new mitophagy reporter mouse line to examine how disuse affected mitophagy activity in skeletal muscles. Mice expressing tandem mCherry-EGFP proteins on mitochondria were then used to monitor the dynamics of mitophagy activity. The reporter mice demonstrated enhanced mitophagy activity and increased ROS production in atrophic soleus muscles following a 14-day hindlimb immobilization. Results also showed an increased expression of multiple mitophagy genes, including Bnip3, Bnip3l, and Park2. Our findings thus conclude that disuse enhances mitophagy activity and ROS production in atrophic skeletal muscles and suggests that mitophagy is a potential therapeutic target for disuse-induced muscle atrophy.
    Keywords:  ROS; disuse-induced muscle atrophy; hindlimb immobilization; mitochondria; mitophagy
    DOI:  https://doi.org/10.1002/jcp.30404
  5. Exp Eye Res. 2021 Oct 14. pii: S0014-4835(21)00359-6. [Epub ahead of print] 108793
    Quantifying inter-organelle membrane contact sites using proximity ligation assay in fixed optic nerve sections.
    Jared Ching, Andrew Osborne, Richard Eva, Julien Prudent, Patrick Yu-Wai-Man.
      Membrane contact sites (MCS) play crucial roles in cell physiology with dysfunction in several MCS proteins being linked with neurological and optic nerve diseases. Although there have been significant advances in imaging these interactions over the past two decades with advanced electron microscopy techniques, super-resolution imaging and proximity-dependent fluorescent reporters, a technique to observe and quantify MCS in mammalian optic nerve tissues has not yet been reported. We demonstrate for the first time that proximity ligation assay (PLA), a technique already used in mammalian cell lines, can be used as an efficient method of quantifying inter-organelle contact sites, namely mitochondria-endoplasmic reticulum (ER) and mitochondria-late-endosomes, in mammalian optic nerve tissues treated with adeno-associated virus (AAV) gene therapy with wild-type or phosphomimetic (active) protrudin. PLA utilises complementary single-stranded DNA oligomers bound to secondary antibodies that hybridise and complete a circular piece of DNA when the primary antibodies of interest interact. These interactions can be detected by amplifying the circular DNA and adding fluorescent probes. We show that PLA is a useful method that can be used to quantify MCS in optic nerve tissues. We have found that upregulation of protrudin with gene therapy significantly increases the number of mitochondria-ER and mitochondria-Rab7-late endosomes contact sites in optic nerves.
    Keywords:  Membrane contact sites; Optic nerve; Protrudin; Proximity ligation assay; Retinal ganglion cells
    DOI:  https://doi.org/10.1016/j.exer.2021.108793
  6. Exp Cell Res. 2021 Oct 13. pii: S0014-4827(21)00420-1. [Epub ahead of print] 112866
    Alleviation of CCCP injury by Augmenter of liver regeneration via the PINK1/Parkin pathway-dependent mitophagy.
    Jing Zhang, Sili Chen, Ye Li, Weichun Xiao, Wei An.
      The occurrence of liver diseases is attributed to mitochondrial damage. Mitophagy selectively removes dysfunctional mitochondria, thereby preserving mitochondrial function. Augmenter of liver regeneration (ALR) protects the mitochondria from injury. However, whether ALR protection is associated with mitophagy remains unclear. In this study, mitochondrial damage was induced by carbonyl cyanide 3-chlorophenylhydrazone (CCCP), and long form ALR (lfRNA) -mediated protection against this damage was investigated. Treatment of HepG2 cells with CCCP elevated the level of intracellular ROS, inhibited ATP production, and increased the mitochondrial membrane potential and cell apoptotic rate. However, in lfALR-transfected cells, CCCP-induced cell injury was clearly alleviated, the apoptosis and ROS levels clearly declined, and the ATP production was significantly enhanced as compared with that in vector-Tx cells. Furthermore, lfALR overexpression promoted autophagy and mitophagy via a PINK1/Parkin-dependent pathway, whereas knockdown of ALR suppressed mitophagy. In lfALR-transfected cells, the phosphorylation of AKT was decreased, thus, downregulating the phosphorylation of the transcription factor FOXO3a at Ser315. In contrast, the phosphorylation of AMPK was enhanced, thereby upregulating the phosphorylation of FOXO3a at Ser413. Consequently, FOXO3a's nuclear translocation and binding to the promoter region of PINK1 was enhanced, and the accumulation of PINK1/Parkin in mitochondria increased. Meanwhile, short form ALR (sfALR) also increased PINK1 expression through FOXO3a with the similar pathway to lfALR. In conclusion, our data suggest a novel mechanism through which both lfALR and sfALR protect mitochondria by promoting PINK1/Parkin-dependent mitophagy through FOXO3a activation.
    Keywords:  Augmenter of liver regeneration; FOXO3a; Mitophagy; PINK1/Parkin
    DOI:  https://doi.org/10.1016/j.yexcr.2021.112866
  7. J Cell Sci. 2021 Oct 21. pii: jcs.259091. [Epub ahead of print]
    A three-organelle complex made by wrappER contacts with peroxisome and mitochondria responds to liver lipid flux changes.
    Nicolò Ilacqua, Irene Anastasia, Andrea Raimondi, Philippe Lemieux, Thomas Q de Aguiar Vallim, Katalin Toth, Eugene V Koonin, Luca Pellegrini.
      Hepatic lipid homeostasis depends on intracellular pathways that respire fatty acid (FA) in peroxisomes and mitochondria and on systemic pathways that secrete FA into the bloodstream, either free or condensed in very-low-density lipoprotein (VLDL) triglycerides. These systemic and intracellular pathways are interdependent, but it is unclear whether and how they integrate into a single cellular circuit. Here, we report that mouse liver wrappER, a distinct ER compartment with apparent FA- and VLDL-secretion functions, connects peroxisomes and mitochondria. Correlative light electron microscopy, quantitative serial section electron tomography, and 3D organelle reconstruction analysis show that the number of peroxisome-wrappER-mitochondria complexes changes throughout fasting-to-feeding transitions and doubles when VLDL synthesis stops following acute genetic ablation of Mttp in the liver. Quantitative proteomic analysis of peroxisome-wrappER-mitochondria complex-enriched fractions indicates that the loss of Mttp upregulates global FA β-oxidation, thereby integrating the dynamics of this three-organelle association into hepatic FA flux responses. Therefore, liver lipid homeostasis occurs through the convergence of systemic and intracellular FA-elimination pathways in the peroxisome-wrappER-mitochondria complex.
    Keywords:  Fatty acid; Inter-organelle contacts; Liver lipid homeostasis; Mitochondria; Peroxisome; WrappER
    DOI:  https://doi.org/10.1242/jcs.259091
  8. Front Aging Neurosci. 2021 ;13 755665
    Exercise-Induced Benefits for Alzheimer's Disease by Stimulating Mitophagy and Improving Mitochondrial Function.
    Jiling Liang, Cenyi Wang, Hu Zhang, Jielun Huang, Juying Xie, Ning Chen.
      Neurons are highly specialized post-mitotic cells that are inherently dependent on mitochondria due to their higher bioenergetic demand. Mitochondrial dysfunction is closely associated with a variety of aging-related neurological disorders, such as Alzheimer's disease (AD), and the accumulation of dysfunctional and superfluous mitochondria has been reported as an early stage that significantly facilitates the progression of AD. Mitochondrial damage causes bioenergetic deficiency, intracellular calcium imbalance and oxidative stress, thereby aggravating β-amyloid (Aβ) accumulation and Tau hyperphosphorylation, and further leading to cognitive decline and memory loss. Although there is an intricate parallel relationship between mitochondrial dysfunction and AD, their triggering factors, such as Aβ aggregation and hyperphosphorylated Tau protein and action time, are still unclear. Moreover, many studies have confirmed abnormal mitochondrial biosynthesis, dynamics and functions will present once the mitochondrial quality control is impaired, thus leading to aggravated AD pathological changes. Accumulating evidence shows beneficial effects of appropriate exercise on improved mitophagy and mitochondrial function to promote mitochondrial plasticity, reduce oxidative stress, enhance cognitive capacity and reduce the risks of cognitive impairment and dementia in later life. Therefore, stimulating mitophagy and optimizing mitochondrial function through exercise may forestall the neurodegenerative process of AD.
    Keywords:  Alzheimer’s disease; exercise; mitochondrial quality control; mitophagy; oxidative stress
    DOI:  https://doi.org/10.3389/fnagi.2021.755665
  9. Int Immunopharmacol. 2021 Oct 13. pii: S1567-5769(21)00866-3. [Epub ahead of print]101(Pt B): 108230
    Apelin-13-Mediated AMPK ameliorates endothelial barrier dysfunction in acute lung injury mice via improvement of mitochondrial function and autophagy.
    Xiaoxia Kong, Daopeng Lin, Liling Lu, Lidan Lin, Hongyu Zhang, Hailin Zhang.
      Maintaining the pulmonary endothelial barrier that prevents the exudation of inflammatory factors and proteins is the key to the treatment of acute lung injury (ALI). Apelin-13 plays an important role in vascular diseases; however, the protective effects of Apelin-13 on ALI with pulmonary endothelial barrier are unknown. Therefore, mice and human umbilical vein endothelial cells (HUVECs) were injured by LPS following Apelin-13 administration. ALI mice showed reduced pulmonary vascular permeability, adhesion junction, mitochondrial function, mitochondrial biogenesis, and autophagy compared to the control group. Apelin-13 administration in ALI mice ameliorated LPS-induced lung injury, pulmonary vascular permeability, mitochondrial function, and promoted autophagic flux in mice and HUVECs. However, the effect of Apelin-13 was reduced after AMPK inhibition using Compound C. These data suggest that Apelin-13 ameliorates pulmonary vascular permeability in mice with ALI induced by LPS, which may be related to enhanced phosphorylation of AMPK to regulate mitochondrial function and autophagy.
    Keywords:  AMPK; Acute lung injury; Apelin-13; Autophagy; Mitochondrial biogenesis
    DOI:  https://doi.org/10.1016/j.intimp.2021.108230
  10. Biomolecules. 2021 Oct 13. pii: 1508. [Epub ahead of print]11(10):
    Mitochondrial Dysfunction, Protein Misfolding and Neuroinflammation in Parkinson's Disease: Roads to Biomarker Discovery.
    Anna Picca, Flora Guerra, Riccardo Calvani, Roberta Romano, Hélio José Coelho-Júnior, Cecilia Bucci, Emanuele Marzetti.
      Parkinson's Disease (PD) is a highly prevalent neurodegenerative disease among older adults. PD neuropathology is marked by the progressive loss of the dopaminergic neurons of the substantia nigra pars compacta and the widespread accumulation of misfolded intracellular α-synuclein (α-syn). Genetic mutations and post-translational modifications, such as α-syn phosphorylation, have been identified among the multiple factors supporting α-syn accrual during PD. A decline in the clearance capacity of the ubiquitin-proteasome and the autophagy-lysosomal systems, together with mitochondrial dysfunction, have been indicated as major pathophysiological mechanisms of PD neurodegeneration. The accrual of misfolded α-syn aggregates into soluble oligomers, and the generation of insoluble fibrils composing the core of intraneuronal Lewy bodies and Lewy neurites observed during PD neurodegeneration, are ignited by the overproduction of reactive oxygen species (ROS). The ROS activate the α-syn aggregation cascade and, together with the Lewy bodies, promote neurodegeneration. However, the molecular pathways underlying the dynamic evolution of PD remain undeciphered. These gaps in knowledge, together with the clinical heterogeneity of PD, have hampered the identification of the biomarkers that may be used to assist in diagnosis, treatment monitoring, and prognostication. Herein, we illustrate the main pathways involved in PD pathogenesis and discuss their possible exploitation for biomarker discovery.
    Keywords:  DAMPs; alpha-synuclein; beta-amyloid (Aβ); cytokine; extracellular vesicles; inflammation; mitochondrial-derived vesicles; mitophagy; neurofilaments light chain (NfL); p-tau pathology
    DOI:  https://doi.org/10.3390/biom11101508
  11. Autophagy. 2021 Oct 18. 1-16
    Full-coverage regulations of autophagy by ROS: from induction to maturation.
    Jing Zhou, Xin-Yu Li, Yu-Jia Liu, Ji Feng, Yong Wu, Han-Ming Shen, Guo-Dong Lu.
      Macroautophagy/autophagy is an evolutionarily well-conserved recycling process in response to stress conditions, including a burst of reactive oxygen species (ROS) production. High level of ROS attack key cellular macromolecules. Protein cysteinyl thiols or non-protein thiols as the major redox-sensitive targets thus constitute the first-line defense. Autophagy is unique, because it removes not only oxidized/damaged proteins but also bulky ROS-generating organelles (such as mitochondria and peroxisome) to restrict further ROS production. The oxidative regulations of autophagy occur in all processes of autophagy, from induction, phagophore nucleation, phagophore expansion, autophagosome maturation, cargo delivery to the lysosome, and finally to degradation of the cargo and recycling of the products, as well as autophagy gene transcription. Mechanically, these regulations are achieved through direct or indirect manners. Direct thiol oxidation of key proteins such as ATG4, ATM and TFEB are responsible for specific regulations in phagophore expansion, cargo recognition and autophagy gene transcription, respectively. Meanwhile, oxidation of certain redox-sensitive chaperone-like proteins (e.g. PRDX family members and PARK7) may impair a nonspecifically local reducing environment in the phagophore membrane, and influence BECN1-involved phagophore nucleation and mitophagy recognition. However, ROS do exhibit some inhibitory effects on autophagy through direct oxidation of key autophagy regulators such as ATG3, ATG7 and SENP3 proteins. SQSTM1 provides an alternative antioxidant mechanism when autophagy is unavailable or impaired. However, it is yet to be unraveled how cells evolve to equip proteins with different redox susceptibility and in their correct subcellular positions, and how cells fine-tune autophagy machinery in response to different levels of ROS.Abbreviations: AKT1/PKB: AKT serine/threonine kinase 1; AMPK: AMP-activated protein kinase; ATG: autophagy related; ATM: ATM serine/threonine kinase; BAX: BCL2 associated X, apoptosis regulator; BECN1: beclin 1; BH3: BCL2-homology-3; CAV1: caveolin 1; CCCP: carbonyl cyanide m-chlorophenylhydrazone; CTSB: cathepsin B; CTSL: cathepsin L; DAPK: death associated protein kinase; ER: endoplasmic reticulum; ETC: electron transport chain; GSH: glutathione; GSTP1: glutathione S-transferase pi 1; H2O2: hydrogen peroxide; HK2: hexokinase 2; KEAP1: kelch like ECH associated protein 1; MAMs: mitochondria-associated ER membranes; MAP1LC3B/LC3: microtubule associated protein 1 light chain 3 beta; MAPK8/JNK1: mitogen-activated protein kinase 8; MAP3K5/ASK1: mitogen-activated protein kinase kinase kinase 5; MCOLN1: mucolipin 1; MMP: mitochondrial membrane potential; MTOR: mechanistic target of rapamycin kinase; NFE2L2/NRF2: nuclear factor, erythroid 2 like 2; NFKB1: nuclear factor kappa B subunit 1; NOX: NADPH oxidase; O2-: superoxide radical anion; p-Ub: phosphorylated Ub; PARK7/DJ-1: Parkinsonism associated deglycase; PE: phosphatidylethanolamine; PEX5: peroxisomal biogenesis factor 5; PINK1: PTEN induced kinase 1; PPP3CA/calcineurin: protein phosphatase 3 catalytic subunit beta; PRDX: peroxiredoxin; PRKAA1: protein kinase AMP-activated catalytic subunit alpha 1; PRKD/PKD: protein kinase D; PRKN/parkin: parkin RBR E3 ubiquitin protein ligase; PtdIns3K: class III phosphatidylinositol 3-kinase; PtdIns3P: phosphatidylinositol-3-phosphate; PTEN: phosphatase and tensin homolog; ROS: reactive oxygen species; SENP3: SUMO specific peptidase 3; SIRT1: sirtuin 1; SOD1: superoxide dismutase 1; SQSTM1/p62: sequestosome 1; SUMO: small ubiquitin like modifier; TFEB: transcription factor EB; TRAF6: TNF receptor associated factor 6; TSC2: TSC complex subunit 2; TXN: thioredoxin; TXNRD1: thioredoxin reductase 1; TXNIP: thioredoxin interacting protein; Ub: ubiquitin; ULK1: unc-51 like autophagy activating kinase 1.
    Keywords:  ATGs; ROS; autophagy; oxidative regulation; protein thiols
    DOI:  https://doi.org/10.1080/15548627.2021.1984656
  12. Nat Protoc. 2021 Oct 22.
    Quantification of organelle contact sites by split-GFP-based contact site sensors (SPLICS) in living cells.
    Tito Calì, Marisa Brini.
      Membrane contact sites between organelles are essential for maintaining cellular homeostasis, which requires the continuous exchange of signaling molecules, ions, nutrients and lipids. Alterations of different contact sites are associated with a wide spectrum of human diseases. However, visualizing and quantifying these contact sites remains a challenge. This protocol describes the use of split-GFP-based contact site sensors (SPLICS) in microscopy applications for mapping organelle contact sites both in fixed and living cells. SPLICS sensors are engineered to express equimolar amounts of the organelle-targeted nonfluorescent β11 and GFP1-10 portions of the split-GFP protein in a single vector, and are capable of reconstituting fluorescence when two opposing membranes come into proximity. Reconstitution will occur only over the cell volume at defined contact sites resulting in a bright signal that can be detected easily and quantified automatically with specific custom-made plugins. The use of minimal targeting sequences facilitates targeting specificity and membrane coverage, avoiding artifacts due to full-length fusion protein overexpression and, thus, possible perturbations of the cell's physiology. SPLICS sensors engineered to simultaneously detect multiple contact sites within the same cell have been generated by exploiting the ability of the β11 GFP fragment to reconstitute different color-shifted variants of the GFP1-10 fragment. Here, we describe a detailed protocol to set up SPLICS expression in living cells (2-3 d), detection and acquisition (1 d), and automated quantification with custom plugins (1-2 d). We also advise on construct design and characterization for novel organelle contacts.
    DOI:  https://doi.org/10.1038/s41596-021-00614-1
  13. Cells. 2021 Oct 02. pii: 2639. [Epub ahead of print]10(10):
    New Insights into Molecular Mechanisms Mediating Adaptation to Exercise; A Review Focusing on Mitochondrial Biogenesis, Mitochondrial Function, Mitophagy and Autophagy.
    Fiona Louise Roberts, Greg Robert Markby.
      Exercise itself is fundamental for good health, and when practiced regularly confers a myriad of metabolic benefits in a range of tissues. These benefits are mediated by a range of adaptive responses in a coordinated, multi-organ manner. The continued understanding of the molecular mechanisms of action which confer beneficial effects of exercise on the body will identify more specific pathways which can be manipulated by therapeutic intervention in order to prevent or treat various metabolism-associated diseases. This is particularly important as exercise is not an available option to all and so novel methods must be identified to confer the beneficial effects of exercise in a therapeutic manner. This review will focus on key emerging molecular mechanisms of mitochondrial biogenesis, autophagy and mitophagy in selected, highly metabolic tissues, describing their regulation and contribution to beneficial adaptations to exercise.
    Keywords:  autophagy; exercise; mitophagy and mitochondrial biogenesis; molecular signaling
    DOI:  https://doi.org/10.3390/cells10102639
  14. Cell Rep. 2021 Oct 19. pii: S2211-1247(21)01312-7. [Epub ahead of print]37(3): 109848
    Lysosomal Zn2+ release triggers rapid, mitochondria-mediated, non-apoptotic cell death in metastatic melanoma.
    Wanlu Du, Mingxue Gu, Meiqin Hu, Prateeksunder Pinchi, Wei Chen, Michael Ryan, Timothy Nold, Ahmed Bannaga, Haoxing Xu.
      During tumor progression, lysosome function is often maladaptively upregulated to match the high energy demand required for cancer cell hyper-proliferation and invasion. Here, we report that mucolipin TRP channel 1 (TRPML1), a lysosomal Ca2+ and Zn2+ release channel that regulates multiple aspects of lysosome function, is dramatically upregulated in metastatic melanoma cells compared with normal cells. TRPML-specific synthetic agonists (ML-SAs) are sufficient to induce rapid (within hours) lysosomal Zn2+-dependent necrotic cell death in metastatic melanoma cells while completely sparing normal cells. ML-SA-caused mitochondria swelling and dysfunction lead to cellular ATP depletion. While pharmacological inhibition or genetic silencing of TRPML1 in metastatic melanoma cells prevents such cell death, overexpression of TRPML1 in normal cells confers ML-SA vulnerability. In the melanoma mouse models, ML-SAs exhibit potent in vivo efficacy of suppressing tumor progression. Hence, targeting maladaptively upregulated lysosome machinery can selectively eradicate metastatic tumor cells in vitro and in vivo.
    Keywords:  ML-SAs; ML-SIs; TRPML1; Zn(2+); cell death; lysosome; metastatic melanoma; mitochondria; small molecule
    DOI:  https://doi.org/10.1016/j.celrep.2021.109848
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