bims-mitlys Biomed News
on Mitochondria and Lysosomes
Issue of 2021‒07‒11
five papers selected by
Nicoletta Plotegher
University of Padua

  1. Methods Cell Biol. 2021 ;pii: S0091-679X(20)30212-0. [Epub ahead of print]164 113-118
      Mitophagy is an evolutionally conserved cellular process that eliminates dysfunctional and excess mitochondria, thereby facilitating mitochondrial quality control and metabolic recycling. In addition, mitophagy is essential for cellular homeostasis and tissue development, and mitophagic dysfunction is related to various pathologies including neurodegenerative diseases and cancer. Thus, accurate quantitative measurement of mitophagy is one of the hot topics in the field of mitochondrial research. Fluorescence microscopical technology, one of the most widely used technologies at present, can well explain the occurrence and activity of mitophagy. Here, we introduce in detail an experimental method for the immunofluorescence-based quantitativ determination of mitophagy, which not only servers the in-depth study of mitochondrial homeostasis regulation, but also allows for the analyzing mitochondrial autophagy pathologies such as aging, neurodegenerative diseases and cancer.
    Keywords:  Detection; Fluorescence microscope; Method; MitoTracker; Mitophagy
  2. Autophagy. 2021 Jul 07. 1-18
      There is increasing evidence that mitophagy, a specialized form of autophagy to degrade and clear long-lived or damaged mitochondria, is impaired in aging and age-related disease. Previous study has demonstrated the obesity-exposed oocytes accumulate and transmit damaged mitochondria due to an inability to activate mitophagy. However, it remains unknown whether mitophagy functions in oocyte and what's the regulatory mechanism in oocyte aging. In the study, when fully grown oocytes were treated with CCCP, an uncoupling agent to induce mitophagy, we found the activation of the PRKN-mediated mitophagy pathway accompanied the blockage of meiosis at metaphase I stage. Our result then demonstrated its association with the decreased activity of RAB7 and all the observed defects in CCCP treated oocytes could be effectively rescued by microinjection of mRNA encoding active RAB7Q67L or treatment with the RAB7 activator ML098. Further study indicated PRKN protein level as a rate-limiting factor to facilitate degradation of RAB7 and its GEF (guanine nucleotide exchange factor) complex CCZ1-MON1 through the ubiquitin-proteasome system. In GV oocytes collected during ovarian aging, we found the age-related increase of PINK1 and PRKN proteins and a significant decrease of RAB7 which resulted in defects of mitophagosome formation and the accumulation of damaged mitochondria. The age-related retardation of female fertility was improved after in vivo treatment of ML098. Thus, RAB7 activity is required to maintain the balance between mitophagy and chromosome stability and RAB7 activator is a good candidate to ameliorate age-related deterioration of oocyte quality.Abbreviations: ATG9: autophagy related 9A; ATP: adenosine triphosphate; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CCCP: carbonyl cyanide 3-chlorophenylhydrazone; CCZ1: CCZ1 vacuolar protein trafficking and biogenesis associated; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GAPs: GTPase-activating proteins; GEF: guanine nucleotide exchange factor; GV: germinal vesicle; GVBD: germinal vesicle breakdown; LAMP1: lysosomal-associated membrane protein 1; MI: metaphase I stage of meiosis; MII: metaphase II stage of meiosis; Mito: MitoTracker; mtDNA: mitochondrial DNA; MON1: MON1 homolog, secretory trafficking associated; OPTN: optineurin; PINK1: PTEN induced putative kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; RAB7: RAB7, member RAS oncogene family; ROS: reactive oxygen species; TEM: transmission electron microscopy; TOMM20/TOM20: translocase of outer mitochondrial membrane 20; TUBB: tubulin, beta; UB: ubiquitin.
    Keywords:  Aging; PRKN; RAB7; meiosis; mitophagy; oocyte
  3. Methods Cell Biol. 2021 ;pii: S0091-679X(21)00014-5. [Epub ahead of print]164 95-112
      In the perspective to evaluate the toxicity of drug candidates or the exploration of intracellular signaling pathways of cell stress response and pathophysiological conditions, we propose to evaluate cell death, autophagy, mitochondrial network and energetic metabolism by a series of optimized joint protocols for neonatal primary rat cardiomyocytes or H9c2 cardiac cell line in 96 well microtiter plates. We used Digitoxigenin and Digoxin, two cardiac glycosides, and Rapamycin as control drugs, for inhibition of oxidative stress-induced cell death and autophagy induction, respectively.
    Keywords:  Autophagy; Cell death; Cytotoxicity; Metabolism; Mitochondria; Viability
  4. Cell Death Dis. 2021 Jul 03. 12(7): 671
      The balanced functionality of cellular proteostatic modules is central to both proteome stability and mitochondrial physiology; thus, the age-related decline of proteostasis also triggers mitochondrial dysfunction, which marks multiple degenerative disorders. Non-functional mitochondria are removed by mitophagy, including Parkin/Pink1-mediated mitophagy. A common feature of neuronal or muscle degenerative diseases, is the accumulation of damaged mitochondria due to disrupted mitophagy rates. Here, we exploit Drosophila as a model organism to investigate the functional role of Parkin/Pink1 in regulating mitophagy and proteostatic responses, as well as in suppressing degenerative phenotypes at the whole organism level. We found that Parkin or Pink1 knock down in young flies modulated proteostatic components in a tissue-dependent manner, increased cell oxidative load, and suppressed mitophagy in neuronal and muscle tissues, causing mitochondrial aggregation and neuromuscular degeneration. Concomitant to Parkin or Pink1 knock down cncC/Nrf2 overexpression, induced the proteostasis network, suppressed oxidative stress, restored mitochondrial function, and elevated mitophagy rates in flies' tissues; it also, largely rescued Parkin or Pink1 knock down-mediated neuromuscular degenerative phenotypes. Our in vivo findings highlight the critical role of the Parkin/Pink1 pathway in mitophagy, and support the therapeutic potency of Nrf2 (a druggable pathway) activation in age-related degenerative diseases.
  5. Cancer Cell Int. 2021 Jul 05. 21(1): 350
      The mitochondrion is an organelle that plays a vital role in energy production, cytoplasmic protein degradation and cell death. Mitophagy is an autophagic procedure that specifically clears damaged mitochondria and maintains its homeostasis. Emerging evidence indicates that mitophagy is involved in many physiological processes, including cellular homeostasis, cellular differentiation and nerve protection. In this review, we describe the regulatory mechanisms of mitophagy in mammals and yeasts and highlight the recent advances relevant to its function in carcinogenesis and drug resistance. Finally, a section has been dedicated to describing the role of mitophagy in anticancer therapeutics, which is a new frontier that offers a precise and promising strategy.
    Keywords:  Anticancer therapeutics; Carcinogenesis; Drug resistance; Mechanism; Mitophagy