bims-mitmed Biomed News
on Mitochondrial medicine
Issue of 2024‒06‒16
twenty-one papers selected by
Dario Brunetti, Fondazione IRCCS Istituto Neurologico 
  1. MTFP1 controls mitochondrial fusion to regulate inner membrane quality control and maintain mtDNA levels.
  2. Nanobiopsy investigation of the subcellular mtDNA heteroplasmy in human tissues.
  3. SS-31 treatment ameliorates cardiac mitochondrial morphology and defective mitophagy in a murine model of Barth syndrome.
  4. Evidence for sodium valproate toxicity in mitochondrial diseases: a systematic analysis.
  5. Mitochondrial membrane synthesis, remodelling and cellular trafficking.
  6. Aberrant ER-mitochondria communication is a common pathomechanism in mitochondrial disease.
  7. Unique architectural features of mammalian mitochondrial protein synthesis.
  8. Reproductive regulation of the mitochondrial stress response in Caenorhabditis elegans.
  9. Hematologic Manifestations in Primary Mitochondrial Diseases.
  10. Role of Yme1 in mitochondrial protein homeostasis: from regulation of protein import, OXPHOS function to lipid synthesis and mitochondrial dynamics.
  11. Privileged proteins with a second residence: dual targeting and conditional re-routing of mitochondrial proteins.
  12. Metabolic inflexibility promotes mitochondrial health during liver regeneration.
  13. Electron transport chain inhibition increases cellular dependence on purine transport and salvage.
  14. Molecular mechanism of the ischemia-induced regulatory switch in mammalian complex I.
  15. A computational study to assess the pathogenicity of single or combinations of missense variants on respiratory complex I.
  16. C16ORF70/Mytho promotes healthy ageing in C. elegans and prevents cellular senescence in mammals.
  17. Efficient site-specific integration of large genes in mammalian cells via continuously evolved recombinases and prime editing.
  18. Release of CD36-associated cell-free mitochondrial DNA and RNA as a hallmark of space environment response.
  19. High-Resolution Fluorespirometry to Assess Dynamic Changes in Mitochondrial Membrane Potential in Human Immune Cells.
  20. Protocol for Seahorse analysis of ex vivo mouse brown and white adipose tissues.
  21. Inborn Errors of Metabolism and Pregnancy.
  1. Cell. 2024 Jun 05. pii: S0092-8674(24)00526-9. [Epub ahead of print]
    MTFP1 controls mitochondrial fusion to regulate inner membrane quality control and maintain mtDNA levels.
    Luis Carlos Tábara, Stephen P Burr, Michele Frison, Suvagata R Chowdhury, Vincent Paupe, Yu Nie, Mark Johnson, Jara Villar-Azpillaga, Filipa Viegas, Mayuko Segawa, Hanish Anand, Kasparas Petkevicius, Patrick F Chinnery, Julien Prudent.
      Mitochondrial dynamics play a critical role in cell fate decisions and in controlling mtDNA levels and distribution. However, the molecular mechanisms linking mitochondrial membrane remodeling and quality control to mtDNA copy number (CN) regulation remain elusive. Here, we demonstrate that the inner mitochondrial membrane (IMM) protein mitochondrial fission process 1 (MTFP1) negatively regulates IMM fusion. Moreover, manipulation of mitochondrial fusion through the regulation of MTFP1 levels results in mtDNA CN modulation. Mechanistically, we found that MTFP1 inhibits mitochondrial fusion to isolate and exclude damaged IMM subdomains from the rest of the network. Subsequently, peripheral fission ensures their segregation into small MTFP1-enriched mitochondria (SMEM) that are targeted for degradation in an autophagic-dependent manner. Remarkably, MTFP1-dependent IMM quality control is essential for basal nucleoid recycling and therefore to maintain adequate mtDNA levels within the cell.
    Keywords:  IMM quality control; IMM remodeling; MTFP1; autophagy; fission and fusion; mitochondria; mitochondrial dynamics; mitophagy; mtDNA
    DOI:  https://doi.org/10.1016/j.cell.2024.05.017
  2. Sci Rep. 2024 Jun 14. 14(1): 13789
    Nanobiopsy investigation of the subcellular mtDNA heteroplasmy in human tissues.
    Alexander Bury, Angela Pyle, Amy E Vincent, Paolo Actis, Gavin Hudson.
      Mitochondrial function is critical to continued cellular vitality and is an important contributor to a growing number of human diseases. Mitochondrial dysfunction is typically heterogeneous, mediated through the clonal expansion of mitochondrial DNA (mtDNA) variants in a subset of cells in a given tissue. To date, our understanding of the dynamics of clonal expansion of mtDNA variants has been technically limited to the single cell-level. Here, we report the use of nanobiopsy for subcellular sampling from human tissues, combined with next-generation sequencing to assess subcellular mtDNA mutation load in human tissue from mitochondrial disease patients. The ability to map mitochondrial mutation loads within individual cells of diseased tissue samples will further our understanding of mitochondrial genetic diseases.
    DOI:  https://doi.org/10.1038/s41598-024-64455-0
  3. Sci Rep. 2024 06 13. 14(1): 13655
    SS-31 treatment ameliorates cardiac mitochondrial morphology and defective mitophagy in a murine model of Barth syndrome.
    Silvia Russo, Domenico De Rasmo, Roberta Rossi, Anna Signorile, Simona Lobasso.
      Barth syndrome (BTHS) is a lethal rare genetic disorder, which results in cardiac dysfunction, severe skeletal muscle weakness, immune issues and growth delay. Mutations in the TAFAZZIN gene, which is responsible for the remodeling of the phospholipid cardiolipin (CL), lead to abnormalities in mitochondrial membrane, including alteration of mature CL acyl composition and the presence of monolysocardiolipin (MLCL). The dramatic increase in the MLCL/CL ratio is the hallmark of patients with BTHS, which is associated with mitochondrial bioenergetics dysfunction and altered membrane ultrastructure. There are currently no specific therapies for BTHS. Here, we showed that cardiac mitochondria isolated from TAFAZZIN knockdown (TazKD) mice presented abnormal ultrastructural membrane morphology, accumulation of vacuoles, pro-fission conditions and defective mitophagy. Interestingly, we found that in vivo treatment of TazKD mice with a CL-targeted small peptide (named SS-31) was able to restore mitochondrial morphology in tafazzin-deficient heart by affecting specific proteins involved in dynamic process and mitophagy. This agrees with our previous data showing an improvement in mitochondrial respiratory efficiency associated with increased supercomplex organization in TazKD mice under the same pharmacological treatment. Taken together our findings confirm the beneficial effect of SS-31 in the amelioration of tafazzin-deficient dysfunctional mitochondria in a BTHS animal model.
    DOI:  https://doi.org/10.1038/s41598-024-64368-y
  4. BMJ Neurol Open. 2024 ;6(1): e000650
    Evidence for sodium valproate toxicity in mitochondrial diseases: a systematic analysis.
    Thiloka E Ratnaike, Nour Elkhateeb, Angela Lochmüller, Christopher Gilmartin, Katherine Schon, Rita Horváth, Patrick F Chinnery.
      Background: We aimed to determine whether sodium valproate (VPA) should be contraindicated in all mitochondrial diseases, due to known VPA-induced severe hepatotoxicity in some mitochondrial diseases.Methods: We systematically reviewed the published literature for mitochondrial DNA (mtDNA) and common nuclear genotypes of mitochondrial diseases using PubMed, Ovid Embase, Ovid Medline and MitoPhen databases. We extracted patient-level data from peer-reviewed articles, published until July 2022, using the Human Phenotype Ontology to manually code clinical presentations for 156 patients with genetic diagnoses from 90 publications.
    Results: There were no fatal adverse drug reactions (ADRs) in the mtDNA disease group (35 patients), and only 1 out of 54 patients with a non-POLG mitochondrial disease developed acute liver failure. There were fatal outcomes in 53/102 (52%) POLG VPA-exposed patients who all harboured recessive mutations.
    Conclusions: Our findings confirm the high risk of severe ADRs in any patient with recessive POLG variants irrespective of the phenotype, and therefore recommend that VPA is contraindicated in this group. However, there was limited evidence of toxicity to support a similar recommendation in other genotypes of mitochondrial diseases.
    Keywords:  EPILEPSY; GENETICS; MITOCHONDRIAL DISORDERS; NEUROGENETICS; NEUROPHARMACOLOGY
    DOI:  https://doi.org/10.1136/bmjno-2024-000650
  5. J Inherit Metab Dis. 2024 Jun 14.
    Mitochondrial membrane synthesis, remodelling and cellular trafficking.
    Martina Messina, Frédéric M Vaz, Shamima Rahman.
      Mitochondria are dynamic cellular organelles with complex roles in metabolism and signalling. Primary mitochondrial disorders are a group of approximately 400 monogenic disorders arising from pathogenic genetic variants impacting mitochondrial structure, ultrastructure and/or function. Amongst these disorders, defects of complex lipid biosynthesis, especially of the unique mitochondrial membrane lipid cardiolipin, and membrane biology are an emerging group characterised by clinical heterogeneity, but with recurrent features including cardiomyopathy, encephalopathy, neurodegeneration, neuropathy and 3-methylglutaconic aciduria. This review discusses lipid synthesis in the mitochondrial membrane, the mitochondrial contact site and cristae organising system (MICOS), mitochondrial dynamics and trafficking, and the disorders associated with defects of each of these processes. We highlight overlapping functions of proteins involved in lipid biosynthesis and protein import into the mitochondria, pointing to an overarching coordination and synchronisation of mitochondrial functions. This review also focuses on membrane interactions between mitochondria and other organelles, namely the endoplasmic reticulum, peroxisomes, lysosomes and lipid droplets. We signpost disorders of these membrane interactions that may explain the observation of secondary mitochondrial dysfunction in heterogeneous pathological processes. Disruption of these organellar interactions ultimately impairs cellular homeostasis and organismal health, highlighting the central role of mitochondria in human health and disease.
    Keywords:  MAM; MERC; MICOS; cardiolipin; cell trafficking; mitochondrial lipid biosynthesis; organellar crosstalk; primary mitochondrial disease
    DOI:  https://doi.org/10.1002/jimd.12766
  6. Cell Death Dis. 2024 Jun 10. 15(6): 405
    Aberrant ER-mitochondria communication is a common pathomechanism in mitochondrial disease.
    Patricia Morcillo, Khushbu Kabra, Kevin Velasco, Hector Cordero, Sarah Jennings, Taekyung D Yun, Delfina Larrea, H Orhan Akman, Eric A Schon.
      Genetic mutations causing primary mitochondrial disease (i.e those compromising oxidative phosphorylation [OxPhos]) resulting in reduced bioenergetic output display great variability in their clinical features, but the reason for this is unknown. We hypothesized that disruption of the communication between endoplasmic reticulum (ER) and mitochondria at mitochondria-associated ER membranes (MAM) might play a role in this variability. To test this, we assayed MAM function and ER-mitochondrial communication in OxPhos-deficient cells, including cybrids from patients with selected pathogenic mtDNA mutations. Our results show that each of the various mutations studied indeed altered MAM functions, but notably, each disorder presented with a different MAM "signature". We also found that mitochondrial membrane potential is a key driver of ER-mitochondrial connectivity. Moreover, our findings demonstrate that disruption in ER-mitochondrial communication has consequences for cell survivability that go well beyond that of reduced ATP output. The findings of a "MAM-OxPhos" axis, the role of mitochondrial membrane potential in controlling this process, and the contribution of MAM dysfunction to cell death, reveal a new relationship between mitochondria and the rest of the cell, as well as providing new insights into the diagnosis and treatment of these devastating disorders.
    DOI:  https://doi.org/10.1038/s41419-024-06781-9
  7. Trends Cell Biol. 2024 Jun 08. pii: S0962-8924(24)00097-7. [Epub ahead of print]
    Unique architectural features of mammalian mitochondrial protein synthesis.
    Oliver Rackham, Martin Saurer, Nenad Ban, Aleksandra Filipovska.
      Mitochondria rely on coordinated expression of their own mitochondrial DNA (mtDNA) with that of the nuclear genome for their biogenesis. The bacterial ancestry of mitochondria has given rise to unique and idiosyncratic features of the mtDNA and its expression machinery that can be specific to different organisms. In animals, the mitochondrial protein synthesis machinery has acquired many new components and mechanisms over evolution. These include several new ribosomal proteins, new stop codons and ways to recognise them, and new mechanisms to deliver nascent proteins into the mitochondrial inner membrane. Here we describe the mitochondrial protein synthesis machinery in mammals and its unique mechanisms of action elucidated to date and highlight the technologies poised to reveal the next generation of discoveries in mitochondrial translation.
    Keywords:  RNA; mitochondria; mitochondrial disease; ribosomes; translation
    DOI:  https://doi.org/10.1016/j.tcb.2024.05.001
  8. Cell Rep. 2024 Jun 07. pii: S2211-1247(24)00664-8. [Epub ahead of print]43(6): 114336
    Reproductive regulation of the mitochondrial stress response in Caenorhabditis elegans.
    Nikolaos Charmpilas, Aggeliki Sotiriou, Konstantinos Axarlis, Nektarios Tavernarakis, Thorsten Hoppe.
      Proteome integrity is fundamental for cellular and organismal homeostasis. The mitochondrial unfolded protein response (UPRmt), a key component of the proteostasis network, is activated in a non-cell-autonomous manner in response to mitochondrial stress in distal tissues. However, the importance of inter-tissue communication for UPRmt inducibility under physiological conditions remains elusive. Here, we show that an intact germline is essential for robust UPRmt induction in the Caenorhabditis elegans somatic tissues. A series of nematode mutants with germline defects are unable to respond to genetic or chemical UPRmt inducers. Our genetic analysis suggests that reproductive signals, rather than germline stem cells, are responsible for somatic UPRmt induction. Consistent with this observation, we show that UPRmt is sexually dimorphic, as male nematodes are inherently unresponsive to mitochondrial stress. Our findings highlight a paradigm of germline-somatic communication and suggest that reproductive cessation is a primary cause of age-related UPRmt decline.
    Keywords:  C. elegans; CP: Developmental biology; CP: Molecular biology; aging; germline; mitochondria; proteostasis; unfolded protein response
    DOI:  https://doi.org/10.1016/j.celrep.2024.114336
  9. J Pediatr Hematol Oncol. 2024 Jun 10.
    Hematologic Manifestations in Primary Mitochondrial Diseases.
    Arthavan Selvanathan, Juliana Teo, Bindu Parayil Sankaran.
      Primary mitochondrial disorders (PMDs) are known for their pleiotropic manifestations in humans, affecting almost any organ or system at any time. Hematologic manifestations, such as cytopenias and sideroblastic anemia, occur in 10% to 30% of patients with confirmed PMDs. These can be the initial presenting features or complications that develop over time. Surveillance for these manifestations allows for prompt identification and treatment. This article provides an overview of the pathophysiology underpinning the hematologic effects of mitochondrial dysfunction, discussing the 3 key roles of the mitochondria in hematopoiesis: providing energy for cell differentiation and function, synthesizing heme, and generating iron-sulfur clusters. Subsequently, the diagnosis and management of mitochondrial disorders are discussed, focusing on hematologic manifestations and the specific conditions commonly associated with them. Through this, we aimed to provide a concise point of reference for those considering a mitochondrial cause for a patient's hematologic abnormality, or for those considering a hematologic manifestation in a patient with known or suspected mitochondrial disease.
    DOI:  https://doi.org/10.1097/MPH.0000000000002890
  10. Biochem Soc Trans. 2024 Jun 12. pii: BST20240450. [Epub ahead of print]
    Role of Yme1 in mitochondrial protein homeostasis: from regulation of protein import, OXPHOS function to lipid synthesis and mitochondrial dynamics.
    Kwan Ting Kan, Joel Wilcock, Hui Lu.
      Mitochondria are essential organelles of eukaryotic cells and thus mitochondrial proteome is under constant quality control and remodelling. Yme1 is a multi-functional protein and subunit of the homo-hexametric complex i-AAA proteinase. Yme1 plays vital roles in the regulation of mitochondrial protein homeostasis and mitochondrial plasticity, ranging from substrate degradation to the regulation of protein functions involved in mitochondrial protein biosynthesis, energy production, mitochondrial dynamics, and lipid biosynthesis and signalling. In this mini review, we focus on discussing the current understanding of the roles of Yme1 in mitochondrial protein import via TIM22 and TIM23 pathways, oxidative phosphorylation complex function, as well as mitochondrial lipid biosynthesis and signalling, as well as a brief discussion of the role of Yme1 in modulating mitochondrial dynamics.
    Keywords:  i-AAA proteinase; mitochondrial protein homeostasis; mitochondrial protein import; oxidative phosphorylation complex; protein function
    DOI:  https://doi.org/10.1042/BST20240450
  11. FEBS J. 2024 Jun 10.
    Privileged proteins with a second residence: dual targeting and conditional re-routing of mitochondrial proteins.
    Ophry Pines, Margalit Horwitz, Johannes M Herrmann.
      Almost all mitochondrial proteins are encoded by nuclear genes and synthesized in the cytosol as precursor proteins. Signals in the amino acid sequence of these precursors ensure their targeting and translocation into mitochondria. However, in many cases, only a certain fraction of a specific protein is transported into mitochondria, while the rest either remains in the cytosol or undergoes reverse translocation to the cytosol, and can populate other cellular compartments. This phenomenon is called dual localization which can be instigated by different mechanisms. These include alternative start or stop codons, differential transcripts, and ambiguous or competing targeting sequences. In many cases, dual localization might serve as an economic strategy to reduce the number of required genes; for example, when the same groups of enzymes are required both in mitochondria and chloroplasts or both in mitochondria and the nucleus/cytoplasm. Such cases frequently employ ambiguous targeting sequences to distribute proteins between both organelles. However, alternative localizations can also be used for signaling, for example when non-imported precursors serve as mitophagy signals or when they represent transcription factors in the nucleus to induce the mitochondrial unfolded stress response. This review provides an overview regarding the mechanisms and the physiological consequences of dual targeting.
    Keywords:  dual targeting; mitochondria; protein import; start codon; targeting signals
    DOI:  https://doi.org/10.1111/febs.17191
  12. Science. 2024 Jun 14. 384(6701): eadj4301
    Metabolic inflexibility promotes mitochondrial health during liver regeneration.
    Xun Wang, Cameron J Menezes, Yuemeng Jia, Yi Xiao, Siva Sai Krishna Venigalla, Feng Cai, Meng-Hsiung Hsieh, Wen Gu, Liming Du, Jessica Sudderth, Dohun Kim, Spencer D Shelton, Claire B Llamas, Yu-Hsuan Lin, Min Zhu, Salma Merchant, Divya Bezwada, Sherwin Kelekar, Lauren G Zacharias, Thomas P Mathews, Gerta Hoxhaj, R Max Wynn, Uttam K Tambar, Ralph J DeBerardinis, Hao Zhu, Prashant Mishra.
      Mitochondria are critical for proper organ function and mechanisms to promote mitochondrial health during regeneration would benefit tissue homeostasis. We report that during liver regeneration, proliferation is suppressed in electron transport chain (ETC)-dysfunctional hepatocytes due to an inability to generate acetyl-CoA from peripheral fatty acids through mitochondrial β-oxidation. Alternative modes for acetyl-CoA production from pyruvate or acetate are suppressed in the setting of ETC dysfunction. This metabolic inflexibility forces a dependence on ETC-functional mitochondria and restoring acetyl-CoA production from pyruvate is sufficient to allow ETC-dysfunctional hepatocytes to proliferate. We propose that metabolic inflexibility within hepatocytes can be advantageous by limiting the expansion of ETC-dysfunctional cells.
    DOI:  https://doi.org/10.1126/science.adj4301
  13. Cell Metab. 2024 Jun 07. pii: S1550-4131(24)00190-6. [Epub ahead of print]
    Electron transport chain inhibition increases cellular dependence on purine transport and salvage.
    Zheng Wu, Divya Bezwada, Feng Cai, Robert C Harris, Bookyung Ko, Varun Sondhi, Chunxiao Pan, Hieu S Vu, Phong T Nguyen, Brandon Faubert, Ling Cai, Hongli Chen, Misty Martin-Sandoval, Duyen Do, Wen Gu, Yuanyuan Zhang, Yuannyu Zhang, Bailey Brooks, Sherwin Kelekar, Lauren G Zacharias, K Celeste Oaxaca, Joao S Patricio, Thomas P Mathews, Javier Garcia-Bermudez, Min Ni, Ralph J DeBerardinis.
      Mitochondria house many metabolic pathways required for homeostasis and growth. To explore how human cells respond to mitochondrial dysfunction, we performed metabolomics in fibroblasts from patients with various mitochondrial disorders and cancer cells with electron transport chain (ETC) blockade. These analyses revealed extensive perturbations in purine metabolism, and stable isotope tracing demonstrated that ETC defects suppress de novo purine synthesis while enhancing purine salvage. In human lung cancer, tumors with markers of low oxidative mitochondrial metabolism exhibit enhanced expression of the salvage enzyme hypoxanthine phosphoribosyl transferase 1 (HPRT1) and high levels of the HPRT1 product inosine monophosphate. Mechanistically, ETC blockade activates the pentose phosphate pathway, providing phosphoribosyl diphosphate to drive purine salvage supplied by uptake of extracellular bases. Blocking HPRT1 sensitizes cancer cells to ETC inhibition. These findings demonstrate how cells remodel purine metabolism upon ETC blockade and uncover a new metabolic vulnerability in tumors with low respiration.
    Keywords:  HPRT1; NAD(+):NADH ratio; electron transport chain; metabolomics; purine metabolism; stable isotopes
    DOI:  https://doi.org/10.1016/j.cmet.2024.05.014
  14. Science. 2024 Jun 14. 384(6701): 1247-1253
    Molecular mechanism of the ischemia-induced regulatory switch in mammalian complex I.
    Daniel N Grba, John J Wright, Zhan Yin, William Fisher, Judy Hirst.
      Respiratory complex I is an efficient driver for oxidative phosphorylation in mammalian mitochondria, but its uncontrolled catalysis under challenging conditions leads to oxidative stress and cellular damage. Ischemic conditions switch complex I from rapid, reversible catalysis into a dormant state that protects upon reoxygenation, but the molecular basis for the switch is unknown. We combined precise biochemical definition of complex I catalysis with high-resolution cryo-electron microscopy structures in the phospholipid bilayer of coupled vesicles to reveal the mechanism of the transition into the dormant state, modulated by membrane interactions. By implementing a versatile membrane system to unite structure and function, attributing catalytic and regulatory properties to specific structural states, we define how a conformational switch in complex I controls its physiological roles.
    DOI:  https://doi.org/10.1126/science.ado2075
  15. Int J Biol Macromol. 2024 Jun 11. pii: S0141-8130(24)03891-1. [Epub ahead of print] 133086
    A computational study to assess the pathogenicity of single or combinations of missense variants on respiratory complex I.
    Laura Rigobello, Francesca Lugli, Leonardo Caporali, Alessio Bartocci, Jacopo Fadanni, Francesco Zerbetto, Luisa Iommarini, Valerio Carelli, Anna Maria Ghelli, Francesco Musiani.
      Variants found in the respiratory complex I (CI) subunit genes encoded by mitochondrial DNA can cause severe genetic diseases. However, it is difficult to establish a priori whether a single or a combination of CI variants may impact oxidative phosphorylation. Here we propose a computational approach based on coarse-grained molecular dynamics simulations aimed at investigating new CI variants. One of the primary CI variants associated with the Leber hereditary optic neuropathy (m.14484 T > C/MT-ND6) was used as a test case and was investigated alone or in combination with two additional rare CI variants whose role remains uncertain. We found that the primary variant positioned in the E-channel region, which is fundamental for CI function, stiffens the enzyme dynamics. Moreover, a new mechanism for the transition between π- and α-conformation in the helix carrying the primary variant is proposed. This may have implications for the E-channel opening/closing mechanism. Finally, our findings show that one of the rare variants, located next to the primary one, further worsens the stiffening, while the other rare variant does not affect CI function. This approach may be extended to other variants candidate to exert a pathogenic impact on CI dynamics, or to investigate the interaction of multiple variants.
    Keywords:  Leber's hereditary optic neuropathy; Mitochondria; Molecular dynamics simulations; Pathological variants
    DOI:  https://doi.org/10.1016/j.ijbiomac.2024.133086
  16. J Clin Invest. 2024 Jun 13. pii: e165814. [Epub ahead of print]
    C16ORF70/Mytho promotes healthy ageing in C. elegans and prevents cellular senescence in mammals.
    Anais Franco-Romero, Valeria Morbidoni, Giulia Milan, Roberta Sartori, Jesper Wulff, Vanina Romanello, Andrea Armani, Leonardo Salviati, Maria Conte, Stefano Salvioli, Claudio Franceschi, Viviana Buonomo, Casey O Swoboda, Paolo Grumati, Luca Pannone, Simone Martinelli, Harold Bj Jefferies, Ivan Dikic, Jennifer van der Laan, Filipe Cabreiro, Douglas P Millay, Sharon A Tooze, Eva Trevisson, Marco Sandri.
      The identification of genes that confer either extension of lifespan or accelerate age-related decline was a step forward in understanding the mechanisms of ageing and revealed that it is partially controlled by genetics and transcriptional programs. Here we discovered that the human DNA sequence C16ORF70 encoded for a protein, named MYTHO (Macroautophagy and YouTH Optimizer), which controls life- and health-span. MYTHO protein is conserved from C. elegans to humans and its mRNA was upregulated in aged mice and elderly people. Deletion of the ortholog myt-1 gene in C. elegans dramatically shortened lifespan and decreased animal survival upon exposure to oxidative stress. Mechanistically, MYTHO is required for autophagy likely because it acts as a scaffold that binds WIPI2 and BCAS3 to recruit and assemble the conjugation system at the phagophore, the nascent autophagosome. We conclude that MYTHO is a transcriptionally regulated initiator of autophagy that is central in promoting stress resistance and healthy ageing.
    Keywords:  Aging; Autophagy; Cell biology; Cellular senescence; Skeletal muscle
    DOI:  https://doi.org/10.1172/JCI165814
  17. Nat Biomed Eng. 2024 Jun 10.
    Efficient site-specific integration of large genes in mammalian cells via continuously evolved recombinases and prime editing.
    Smriti Pandey, Xin D Gao, Nicholas A Krasnow, Amber McElroy, Y Allen Tao, Jordyn E Duby, Benjamin J Steinbeck, Julia McCreary, Sarah E Pierce, Jakub Tolar, Torsten B Meissner, Elliot L Chaikof, Mark J Osborn, David R Liu.
      Methods for the targeted integration of genes in mammalian genomes suffer from low programmability, low efficiencies or low specificities. Here we show that phage-assisted continuous evolution enhances prime-editing-assisted site-specific integrase gene editing (PASSIGE), which couples the programmability of prime editing with the ability of recombinases to precisely integrate large DNA cargoes exceeding 10 kilobases. Evolved and engineered Bxb1 recombinase variants (evoBxb1 and eeBxb1) mediated up to 60% donor integration (3.2-fold that of wild-type Bxb1) in human cell lines with pre-installed recombinase landing sites. In single-transfection experiments at safe-harbour and therapeutically relevant sites, PASSIGE with eeBxb1 led to an average targeted-gene-integration efficiencies of 23% (4.2-fold that of wild-type Bxb1). Notably, integration efficiencies exceeded 30% at multiple sites in primary human fibroblasts. PASSIGE with evoBxb1 or eeBxb1 outperformed PASTE (for 'programmable addition via site-specific targeting elements', a method that uses prime editors fused to recombinases) on average by 9.1-fold and 16-fold, respectively. PASSIGE with continuously evolved recombinases is an unusually efficient method for the targeted integration of genes in mammalian cells.
    DOI:  https://doi.org/10.1038/s41551-024-01227-1
  18. Nat Commun. 2024 Jun 11. 15(1): 4814
    Release of CD36-associated cell-free mitochondrial DNA and RNA as a hallmark of space environment response.
    Nailil Husna, Tatsuya Aiba, Shin-Ichiro Fujita, Yoshika Saito, Dai Shiba, Takashi Kudo, Satoru Takahashi, Satoshi Furukawa, Masafumi Muratani.
      A detailed understanding of how spaceflight affects human health is essential for long-term space exploration. Liquid biopsies allow for minimally-invasive multi-omics assessments that can resolve the molecular heterogeneity of internal tissues. Here, we report initial results from the JAXA Cell-Free Epigenome Study, a liquid biopsy study with six astronauts who resided on the International Space Station (ISS) for more than 120 days. Analysis of plasma cell-free RNA (cfRNA) collected before, during, and after spaceflight confirms previously reported mitochondrial dysregulation in space. Screening with 361 cell surface marker antibodies identifies a mitochondrial DNA-enriched fraction associated with the scavenger receptor CD36. RNA-sequencing of the CD36 fraction reveals tissue-enriched RNA species, suggesting the plasma mitochondrial components originated from various tissues. We compare our plasma cfRNA data to mouse plasma cfRNA data from a previous JAXA mission, which had used on-board artificial gravity, and discover a link between microgravity and the observed mitochondrial responses.
    DOI:  https://doi.org/10.1038/s41467-023-41995-z
  19. J Vis Exp. 2024 May 24.
    High-Resolution Fluorespirometry to Assess Dynamic Changes in Mitochondrial Membrane Potential in Human Immune Cells.
    Ana P Valencia, Gavin Pharaoh, Arthur F Brandao, David J Marcinek.
      Peripheral mononuclear cells (PBMCs) exhibit robust changes in mitochondrial respiratory capacity in response to health and disease. While these changes do not always reflect what occurs in other tissues, such as skeletal muscle, these cells are an accessible and valuable source of viable mitochondria from human subjects. PBMCs are exposed to systemic signals that impact their bioenergetic state. Thus, expanding our tools to interrogate mitochondrial metabolism in this population will elucidate mechanisms related to disease progression. Functional assays of mitochondria are often limited to using respiratory outputs following maximal substrate, inhibitor, and uncoupler concentrations to determine the full range of respiratory capacity, which may not be achievable in vivo. The conversion of adenosine diphosphate (ADP) to adenosine triphosphate (ATP) by ATP-synthase results in a decrease in mitochondrial membrane potential (mMP) and an increase in oxygen consumption. To provide a more integrated analysis of mitochondrial dynamics, this article describes the use of high-resolution fluorespirometry to measure the simultaneous response of oxygen consumption and mitochondrial membrane potential (mMP) to physiologically relevant concentrations of ADP. This technique uses tetramethylrhodamine methylester (TMRM) to measure mMP polarization in response to ADP titrations following maximal hyperpolarization with complex I and II substrates. This technique can be used to quantify how changes in health status, such as aging and metabolic disease, affect the sensitivity of mitochondrial response to energy demand in PBMCs, T-cells, and monocytes from human subjects.
    DOI:  https://doi.org/10.3791/66863
  20. STAR Protoc. 2024 Jun 07. pii: S2666-1667(24)00207-7. [Epub ahead of print]5(2): 103042
    Protocol for Seahorse analysis of ex vivo mouse brown and white adipose tissues.
    Fenfen Wang, Phu M Huynh, Yu A An.
      The mitochondrial stress test is a gold-standard approach for assessing adipose tissue physiological functions and pathological changes. Here, we present a protocol for conducting Seahorse assays using ex vivo mouse brown and white adipose depots. We describe steps for rehydrating the cartridge, preparing freshly harvested fat depots, placing them onto an islet capture plate, and incubating them in a non-CO2 incubator. We then detail procedures for adding mitochondrial stressor solutions and conducting the mitochondrial stress test using the Seahorse XFe24 Analyzer. For complete details on the use and execution of this protocol, please refer to An et al.1.
    Keywords:  Cell-based Assays; Health Sciences; Metabolism
    DOI:  https://doi.org/10.1016/j.xpro.2024.103042
  21. Am J Obstet Gynecol MFM. 2024 Jun 11. pii: S2589-9333(24)00125-3. [Epub ahead of print] 101399
    Inborn Errors of Metabolism and Pregnancy.
    Kristen Murphey, Iris Krishna, Hong Li.
      As the diagnosis and treatment of patients with inborn errors of metabolism has improved dramatically over the years, more people with these conditions are surviving into child-bearing years. Given the changes in metabolism throughout pregnancy, this time presents a unique challenge in their care. Overall metabolic shifts in pregnancy go from anabolism to catabolism driven by endocrinologic changes, along with changes in rates of gluconeogenesis, glucose consumption, amino acid transport, protein consumption, and lipid breakdown, result in a complicated metabolic picture. Additionally, maternal inborn errors of metabolism can affect a fetus, as in phenylketonuria, and fetal inborn errors of metabolism can affect the mother, as in certain fatty acid oxidation disorders. Data on these conditions is often very limited. A summary of the current literature, risks associated with pregnancy in inborn errors of metabolism, and suggestions for management of these conditions will be presented.
    Keywords:  Urea cycle disorders; fatty acid oxidation defects; galactosemia; homocystinuria; maple syrup urine disease; maternal phenylketonuria; organic acidemias
    DOI:  https://doi.org/10.1016/j.ajogmf.2024.101399
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