bims-mitdyn Biomed News
on Mitochondrial dynamics: mechanisms
Issue of 2022‒12‒11
twelve papers selected by
Edmond Chan
Queen’s University, School of Medicine
  1. Principles of mitoribosomal small subunit assembly in eukaryotes.
  2. A library of base editors for the precise ablation of all protein-coding genes in the mouse mitochondrial genome.
  3. Mitochondrial cristae architecture protects against mtDNA release and inflammation.
  4. Substitution of PINK1 Gly411 modulates substrate receptivity and turnover.
  5. Regulation of autophagosome biogenesis and mitochondrial bioenergetics by the cholesterol transport protein GRAMD1C.
  6. Mitochondrial succinate dehydrogenase function is essential for sperm motility and male fertility.
  7. PINK1-mediated mitophagy promotes oxidative phosphorylation and redox homeostasis to induce drug-tolerant persister cancer cells.
  8. Essential role of the mitochondrial Na+/Ca2+ exchanger NCLX in mediating PDE2-dependent neuronal survival and learning.
  9. Mitochondrial PARP1 regulates NAD+-dependent poly ADP-ribosylation of mitochondrial nucleoids.
  10. Mitochondrial biology and dysfunction in secondary mitochondrial disease.
  11. Mitolysosome exocytosis: a novel mitochondrial quality control pathway linked with parkinsonism-like symptoms.
  12. Mitochondrial RNA stimulates heat production in young adipocytes to reduce obesity.
  1. Nature. 2022 Dec 08.
    Principles of mitoribosomal small subunit assembly in eukaryotes.
    Nathan J Harper, Chloe Burnside, Sebastian Klinge.
      Mitochondrial ribosomes (mitoribosomes) synthesize proteins encoded within the mitochondrial genome that are assembled into oxidative phosphorylation complexes. Thus, mitoribosome biogenesis is essential for ATP production and cellular metabolism1. Here we used cryo-electron microscopy to determine 9 structures of native yeast and human mitoribosomal small subunit assembly intermediates at resolutions from 2.4 to 3.8 Å, illuminating the mechanistic basis for how GTPases are employed to control early steps of decoding center formation, how initial rRNA folding and processing events are mediated, and how mitoribosomal proteins play active roles during assembly. Furthermore, this series of intermediates from two species with divergent mitoribosomal architecture uncovers both conserved principles and species-specific adaptations that govern the maturation of mitoribosomal small subunits in eukaryotes. By revealing the dynamic interplay between assembly factors, mitoribosomal proteins, and rRNA required to generate functional subunits, our structural analysis provides a vignette for how molecular complexity and diversity can evolve in large ribonucleoprotein assemblies.
    DOI:  https://doi.org/10.1038/s41586-022-05621-0
  2. Nat Biomed Eng. 2022 Dec 05.
    A library of base editors for the precise ablation of all protein-coding genes in the mouse mitochondrial genome.
    Pedro Silva-Pinheiro, Christian D Mutti, Lindsey Van Haute, Christopher A Powell, Pavel A Nash, Keira Turner, Michal Minczuk.
      The development of curative treatments for mitochondrial diseases, which are often caused by mutations in mitochondrial DNA (mtDNA) that impair energy metabolism and other aspects of cellular homoeostasis, is hindered by an incomplete understanding of the underlying biology and a scarcity of cellular and animal models. Here we report the design and application of a library of double-stranded-DNA deaminase-derived cytosine base editors optimized for the precise ablation of every mtDNA protein-coding gene in the mouse mitochondrial genome. We used the library, which we named MitoKO, to produce near-homoplasmic knockout cells in vitro and to generate a mouse knockout with high heteroplasmy levels and no off-target edits. MitoKO should facilitate systematic and comprehensive investigations of mtDNA-related pathways and their impact on organismal homoeostasis, and aid the generation of clinically meaningful in vivo models of mtDNA dysfunction.
    DOI:  https://doi.org/10.1038/s41551-022-00968-1
  3. Cell Rep. 2022 Dec 06. pii: S2211-1247(22)01657-6. [Epub ahead of print]41(10): 111774
    Mitochondrial cristae architecture protects against mtDNA release and inflammation.
    Baiyu He, Huatong Yu, Shanshan Liu, Huayun Wan, Song Fu, Siqi Liu, Jun Yang, Zihan Zhang, Huanwei Huang, Qi Li, Fengchao Wang, Zhaodi Jiang, Qinghua Liu, Hui Jiang.
      Mitochondrial damage causes mitochondrial DNA (mtDNA) release to activate the type I interferon (IFN-I) response via the cGAS-STING pathway. mtDNA-induced inflammation promotes autoimmune- and aging-related degenerative disorders. However, the global picture of inflammation-inducing mitochondrial damages remains obscure. Here, we have performed a mitochondria-targeted CRISPR knockout screen for regulators of the IFN-I response. Strikingly, our screen reveals dozens of hits enriched with key regulators of cristae architecture, including phospholipid cardiolipin and protein complexes such as OPA1, mitochondrial contact site and cristae organization (MICOS), sorting and assembly machinery (SAM), mitochondrial intermembrane space bridging (MIB), prohibitin (PHB), and the F1Fo-ATP synthase. Disrupting these cristae organizers consistently induces mtDNA release and the STING-dependent IFN-I response. Furthermore, knocking out MTX2, a subunit of the SAM complex whose null mutations cause progeria in humans, induces a robust STING-dependent IFN-I response in mouse liver. Taken together, beyond revealing the central role of cristae architecture to prevent mtDNA release and inflammation, our results mechanistically link mitochondrial cristae disorganization and inflammation, two emerging hallmarks of aging and aging-related degenerative diseases.
    Keywords:  CP: Cell biology; CP: Molecular biology; MICOS; Metaxin2; OPA1; SAM; cGAS-STING; cristae architecture; inflammation; mtDNA release; type I interferon response
    DOI:  https://doi.org/10.1016/j.celrep.2022.111774
  4. Autophagy. 2022 Dec 05. 1-22
    Substitution of PINK1 Gly411 modulates substrate receptivity and turnover.
    Fabienne C Fiesel, Dominika Fričová, Caleb S Hayes, Mathew A Coban, Roman Hudec, Jenny M Bredenberg, Benjamin J Broadway, Briana N Markham, Tingxiang Yan, Paige K Boneski, Gabriella Fiorino, Jens O Watzlawik, Xu Hou, Arthur M McCarty, Laura J Lewis-Tuffin, Jun Zhong, Benjamin J Madden, Alban Ordureau, Heeseon An, Andreas Puschmann, Zbigniew K Wszolek, Owen A Ross, J Wade Harper, Thomas R Caulfield, Wolfdieter Springer.
      The ubiquitin (Ub) kinase-ligase pair PINK1-PRKN mediates the degradation of damaged mitochondria by macroautophagy/autophagy (mitophagy). PINK1 surveils mitochondria and upon stress accumulates on the mitochondrial surface where it phosphorylates serine 65 of Ub to activate PRKN and to drive mitochondrial turnover. While loss of either PINK1 or PRKN is genetically linked to Parkinson disease (PD) and activating the pathway seems to have great therapeutic potential, there is no formal proof that stimulation of mitophagy is always beneficial. Here we used biochemical and cell biological methods to study single nucleotide variants in the activation loop of PINK1 to modulate the enzymatic function of this kinase. Structural modeling and in vitro kinase assays were used to investigate the molecular mechanism of the PINK1 variants. In contrast to the PD-linked PINK1G411S mutation that diminishes Ub kinase activity, we found that the PINK1G411A variant significantly boosted Ub phosphorylation beyond levels of PINK1 wild type. This resulted in augmented PRKN activation, mitophagy rates and increased viability after mitochondrial stress in midbrain-derived, gene-edited neurons. Mechanistically, the G411A variant stabilizes the kinase fold of PINK1 and transforms Ub to adopt the preferred, C-terminally retracted conformation for improved substrate turnover. In summary, we identify a critical role of residue 411 for substrate receptivity that may now be exploited for drug discovery to increase the enzymatic function of PINK1. The genetic substitution of Gly411 to Ala increases mitophagy and may be useful to confirm neuroprotection in vivo and might serve as a critical positive control during therapeutic development.Abbreviations: ATP: adenosine triphosphate; CCCP: carbonyl cyanide m-chlorophenyl hydrazone; Ub-CR: ubiquitin with C-terminally retracted tail; CTD: C-terminal domain (of PINK1); ELISA: enzyme-linked immunosorbent assay; HCI: high-content imaging; IB: immunoblot; IF: immunofluorescence; NPC: neuronal precursor cells; MDS: molecular dynamics simulation; PD: Parkinson disease; p-S65-Ub: ubiquitin phosphorylated at Ser65; RMSF: root mean scare fluctuation; TOMM: translocase of outer mitochondrial membrane; TVLN: ubiquitin with T66V and L67N mutation, mimics Ub-CR; Ub: ubiquitin; WT: wild-type.
    Keywords:  Mitophagy; PINK1; PRKN; parkin; parkinson disease; ubiquitin
    DOI:  https://doi.org/10.1080/15548627.2022.2151294
  5. Autophagy. 2022 Dec 05.
    Regulation of autophagosome biogenesis and mitochondrial bioenergetics by the cholesterol transport protein GRAMD1C.
    Chara Charsou, Matthew Yoke Wui Ng, Anne Simonsen.
      Autophagosomes are crucial components of the cellular recycling machinery that form at endoplasmic reticulum (ER)-associated sites. As the autophagosome membrane is largely devoid of transmembrane proteins, autophagosome biogenesis is thought to be largely regulated by lipid transfer and lipid modifications, as well as membrane-associated proteins. While the membrane origin of autophagosomes and their lipid composition are still incompletely understood, previous studies have found the autophagosome membrane to be enriched in unsaturated fatty acids and have little cholesterol, suggesting that cholesterol removal is an integral step during autophagosome biogenesis. In our study, we demonstrate that short term cholesterol depletion leads to a rapid induction of autophagy and identify the ER-localized cholesterol transport protein GRAMD1C as a negative regulator of starvation-induced macroautophagy/autophagy.
    Keywords:  Aster; GRAMD1C; VASt; autophagy; ccRCC; cholesterol
    DOI:  https://doi.org/10.1080/15548627.2022.2155020
  6. iScience. 2022 Dec 22. 25(12): 105573
    Mitochondrial succinate dehydrogenase function is essential for sperm motility and male fertility.
    Rachel M Woodhouse, Natalya Frolows, Guoqiang Wang, Azelle Hawdon, Edmund Heng Kin Wong, Linda C Dansereau, Yingying Su, Liam D Adair, Elizabeth J New, Ashleigh M Philp, Wei Kang Tan, Andrew Philp, Alyson Ashe.
      Mitochondrial health is crucial to sperm quality and male fertility, but the precise role of mitochondria in sperm function remains unclear. SDHA is a component of the succinate dehydrogenase (SDH) complex and plays a critical role in mitochondria. In humans, SDH activity is positively correlated with sperm quality, and mutations in SDHA are associated with Leigh Syndrome. Here we report that the C. elegans SDHA orthologue SDHA-2 is essential for male fertility: sdha-2 mutants produce dramatically fewer offspring due to defective sperm activation and motility, have hyperfused sperm mitochondria, and disrupted redox balance. Similar sperm motility defects in sdha-1 and icl-1 mutant animals suggest an imbalance in metabolites may underlie the fertility defect. Our results demonstrate a role for SDHA-2 in sperm motility and male reproductive health and establish an animal model of SDH deficiency-associated infertility.
    Keywords:  Biological sciences; biochemistry; cell biology; molecular biology
    DOI:  https://doi.org/10.1016/j.isci.2022.105573
  7. Cancer Res. 2022 Dec 08. pii: CAN-22-2370. [Epub ahead of print]
    PINK1-mediated mitophagy promotes oxidative phosphorylation and redox homeostasis to induce drug-tolerant persister cancer cells.
    Yun Li, Hengxing Chen, Xuan Xie, Bing Yang, Xiaojuan Wang, Jingyuan Zhang, Tian Qiao, Jiao Guan, Yuntan Qiu, Yong-Xin Huang, Duanqing Tian, Xinyi Yao, Daning Lu, H Phillip Koeffler, Yin Zhang, Dong Yin.
      The drug-tolerant persister (DTP) state enables cancer cells to evade cytotoxic stress from anti-cancer therapy. However, the mechanisms governing DTP generation remain poorly understood. Here, we observed that lung adenocarcinoma (LUAD) cells and organoids entered a quiescent DTP state to survive MAPK inhibitor treatment. DTP cells following MAPK inhibition underwent a metabolic switch from glycolysis to oxidative phosphorylation (OXPHOS). PTEN-induced kinase 1 (PINK1), a serine/threonine kinase that initiates mitophagy, was upregulated to maintain mitochondrial homeostasis during DTP generation. PINK1-mediated mitophagy supported DTP cell survival and contributed to poor prognosis. Mechanistically, MAPK pathway inhibition resulted in MYC-dependent transcriptional upregulation of PINK1, leading to mitophagy activation. Mitophagy inhibition using either clinically applicable chloroquine or depletion of PINK1 eradicated drug tolerance and allowed complete response to MAPK inhibitors. This study uncovers PINK1-mediated mitophagy as a novel tumor protective mechanism for DTP generation, providing a therapeutic opportunity to eradicate DTP and achieve complete responses.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-22-2370
  8. Cell Rep. 2022 Dec 06. pii: S2211-1247(22)01655-2. [Epub ahead of print]41(10): 111772
    Essential role of the mitochondrial Na+/Ca2+ exchanger NCLX in mediating PDE2-dependent neuronal survival and learning.
    Maya Rozenfeld, Ivana Savic Azoulay, Tsipi Ben Kasus Nissim, Alexandra Stavsky, Moran Melamed, Grace Stutzmann, Michal Hershfinkel, Ora Kofman, Israel Sekler.
      Impaired phosphodiesterase (PDE) function and mitochondrial Ca2+ (i.e., [Ca2+]m) lead to multiple health syndromes by an unknown pathway. Here, we fluorescently monitor robust [Ca2+]m efflux mediated by the mitochondrial Na+/Ca2+ exchanger NCLX in hippocampal neurons sequentially evoked by caffeine and depolarization. Surprisingly, neuronal depolarization-induced Ca2+ transients alone fail to evoke strong [Ca2+]m efflux in wild-type (WT) neurons. However, pre-treatment with the selective PDE2 inhibitor Bay 60-7550 effectively rescues [Ca2+]m efflux similarly to caffeine. Moreover, PDE2 acts by diminishing mitochondrial cAMP, thus promoting NCLX phosphorylation at its PKA site. We find that the protection of neurons against excitotoxic insults, conferred by PDE2 inhibition in WT neurons, is NCLX dependent. Finally, the administration of Bay 60-7550 enhances new object recognition in WT, but not in NCLX knockout (KO), mice. Our results identify a link between PDE and [Ca2+]m signaling that may provide effective therapy for cognitive and ischemic syndromes.
    Keywords:  Bay 60-7550; CP: Neuroscience; NCLX; PDE2; caffeine; mitochondrial Ca(2+) signaling
    DOI:  https://doi.org/10.1016/j.celrep.2022.111772
  9. Exp Mol Med. 2022 Dec 06.
    Mitochondrial PARP1 regulates NAD+-dependent poly ADP-ribosylation of mitochondrial nucleoids.
    Jong-Hyuk Lee, Mansoor Hussain, Edward W Kim, Shang-Jung Cheng, Anthony K L Leung, Nima Borhan Fakouri, Deborah L Croteau, Vilhelm A Bohr.
      PARPs play fundamental roles in multiple DNA damage recognition and repair pathways. Persistent nuclear PARP activation causes cellular NAD+ depletion and exacerbates cellular aging. However, very little is known about mitochondrial PARP (mtPARP) and poly ADP-ribosylation (PARylation). The existence of mtPARP is controversial, and the biological roles of mtPARP-induced mitochondrial PARylation are unclear. Here, we demonstrate the presence of PARP1 and PARylation in purified mitochondria. The addition of the PARP1 substrate NAD+ to isolated mitochondria induced PARylation, which was suppressed by treatment with the inhibitor olaparib. Mitochondrial PARylation was also evaluated by enzymatic labeling of terminal ADP-ribose (ELTA). To further confirm the presence of mtPARP1, we evaluated mitochondrial nucleoid PARylation by ADP ribose-chromatin affinity purification (ADPr-ChAP) and PARP1 chromatin immunoprecipitation (ChIP). We observed that NAD+ stimulated PARylation and TFAM occupancy on the mtDNA regulatory region D-loop, inducing mtDNA transcription. These findings suggest that PARP1 is integrally involved in mitochondrial PARylation and that NAD+-dependent mtPARP1 activity contributes to mtDNA transcriptional regulation.
    DOI:  https://doi.org/10.1038/s12276-022-00894-x
  10. Open Biol. 2022 Dec;12(12): 220274
    Mitochondrial biology and dysfunction in secondary mitochondrial disease.
    Megan J Baker, Jordan J Crameri, David R Thorburn, Ann E Frazier, Diana Stojanovski.
      Mitochondrial diseases are a broad, genetically heterogeneous class of metabolic disorders characterized by deficits in oxidative phosphorylation (OXPHOS). Primary mitochondrial disease (PMD) defines pathologies resulting from mutation of mitochondrial DNA (mtDNA) or nuclear genes affecting either mtDNA expression or the biogenesis and function of the respiratory chain. Secondary mitochondrial disease (SMD) arises due to mutation of nuclear-encoded genes independent of, or indirectly influencing OXPHOS assembly and operation. Despite instances of novel SMD increasing year-on-year, PMD is much more widely discussed in the literature. Indeed, since the implementation of next generation sequencing (NGS) techniques in 2010, many novel mitochondrial disease genes have been identified, approximately half of which are linked to SMD. This review will consolidate existing knowledge of SMDs and outline discrete categories within which to better understand the diversity of SMD phenotypes. By providing context to the biochemical and molecular pathways perturbed in SMD, we hope to further demonstrate the intricacies of SMD pathologies outside of their indirect contribution to mitochondrial energy generation.
    Keywords:  mitochondria; mitochondrial protein import; mitochondrial quality control; secondary mitochondrial disease
    DOI:  https://doi.org/10.1098/rsob.220274
  11. Biochem Soc Trans. 2022 Dec 09. pii: BST20220726. [Epub ahead of print]
    Mitolysosome exocytosis: a novel mitochondrial quality control pathway linked with parkinsonism-like symptoms.
    Feixiang Bao, Lingyan Zhou, Jiahui Xiao, Xingguo Liu.
      Quality control of mitochondria is essential for their homeostasis and function. Light chain 3 (LC3) associated autophagosomes-mediated mitophagy represents a canonical mitochondrial quality control pathway. Alternative quality control processes, such as mitochondrial-derived vesicles (MDVs), have been discovered, but the intact mitochondrial quality control remains unknown. We recently discovered a novel mitolysosome exocytosis mechanism for mitochondrial quality control in flunarizine (FNZ)-induced mitochondria clearance, where autophagosomes are not required, but rather mitochondria are engulfed directly by lysosomes, mediating mitochondrial secretion. As FNZ results in parkinsonism, we propose that excessive mitolysosome exocytosis is the cause.
    Keywords:  extracellular vesicles; lysosome; mitochondria-free; mitochondrial quality control; parkinsonism
    DOI:  https://doi.org/10.1042/BST20220726
  12. Nat Metab. 2022 Dec 06.
    Mitochondrial RNA stimulates heat production in young adipocytes to reduce obesity.
      
    DOI:  https://doi.org/10.1038/s42255-022-00686-7
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