bims-mitdis 2023-04-16 papers

bims-mitdis

Biomed News

on Mitochondrial disorders

Issue of 2023‒04‒16
forty-four papers selected by
Catalina Vasilescu
Helmholz Munich

Mitochondrial Inheritance Following Nuclear Transfer: From Cloned Animals to Patients with Mitochondrial Disease.
Tom40 in cholesterol transport.
Phenotyping mtDNA-related diseases in childhood: a cohort study of 150 patients.
An intermembrane space protein facilitates completion of mitochondrial division in yeast.
Increased mitochondrial surface area and cristae density in the skeletal muscle of strength athletes.
Mitochondrial redox signaling: a key player in aging and disease.
A role for N6-methyldeoxyadenosine in C. elegans mitochondrial genome regulation.
High-Throughput Microscopy Analysis of Mitochondrial Membrane Potential in 2D and 3D Models.
Structural mechanisms of mitochondrial quality control mediated by PINK1 and parkin.
De novo missense variants in RRAGC lead to a fatal mTORopathy of early childhood.
Strategic validation of variants of uncertain significance in ECHS1 genetic testing.
Variant calling and benchmarking in an era of complete human genome sequences.
Comparison of different gene-therapy methods to treat Leber hereditary optic neuropathy in a mouse model.
A retrospective analysis of phosphatase catalytic subunit gene variants in patients with rare disorders identifies novel candidate neurodevelopmental disease genes.
Synergy of uncoupling proteins (1 and 2) with mitochondrial Ca2+ uptake machinery potentiate mitochondrial uncoupling.
PI4P-Containing Vesicles from Golgi Contribute to Mitochondrial Division by Coordinating with Polymerized Actin.
Mitochondrial cardiomyopathy: a fertile field for research.
Arg-tRNA synthetase links inflammatory metabolism to RNA splicing and nuclear trafficking via SRRM2.
Tau Transfer via Extracellular Vesicles Disturbs the Astrocytic Mitochondrial System.
Genome-wide CRISPR screens identify ILF3 as a mediator of mTORC1-dependent amino acid sensing.
Clinical Untargeted Metabolomics as a Functional Screen to Improve Variant Classification.
Physiological and engineered tRNA aminoacylation.
Defective import of mitochondrial metabolic enzyme elicits ectopic metabolic stress.
Evidence supporting the MICU1 occlusion mechanism and against the potentiation model in the mitochondrial calcium uniporter complex.
Cryo-EM structures of mitochondrial ABC transporter ABCB10 in apo and biliverdin-bound form.
Mitochondrial stress response gene Clpp deficiency impairs oocyte competence and deteriorate cyclophosphamide-induced ovarian damage in young mice.
FDXR-associated disease: a challenging differential diagnosis with inflammatory peripheral neuropathy.
A gene pathogenicity tool 'GenePy' identifies missed biallelic diagnoses in the 100,000 Genomes Project.
From recurrent rhabdomyolysis in a young adult to carnitine palmitoyltransferase II deficiency.
Mitochondrial Signaling Pathways Associated with DNA Damage Responses.
IDH3γ functions as a redox switch regulating mitochondrial energy metabolism and contractility in the heart.
Enhanced Mitochondria-SR Tethering Triggers Adaptive Cardiac Muscle Remodeling.
Metabolic regulation of misfolded protein import into mitochondria.
Cardiolipin and OPA1 Team up for Methamphetamine-Induced Locomotor Activity by Promoting Neuronal Mitochondrial Fusion in the Nucleus Accumbens of Mice.
Mitochondrial nanomotion measured by optical microscopy.
Scalable generation of sensory neurons from human pluripotent stem cells.
Denmark's big tent includes postdocs.
Clinical implications of conflicting variant interpretations in the cancer genetics clinic.
The Role of Alternative Mitophagy in Heart Disease.
Clinical and radiological description of 120 pediatric stroke-like episodes.
Advancing Understanding of Inequities in Rare Disease Genomics.
Combined sequencing of genomes and epigenomes.
Using the Reactome Database.
Identifying responders to elamipretide in Barth syndrome: Hierarchical clustering for time series data.

Methods Mol Biol. 2023 ;2647 83-104

Mitochondrial Inheritance Following Nuclear Transfer: From Cloned Animals to Patients with Mitochondrial Disease.

Jörg P Burgstaller, Marcos R Chiaratti.

  Mitochondria are indispensable power plants of eukaryotic cells that also act as a major biochemical hub. As such, mitochondrial dysfunction, which can originate from mutations in the mitochondrial genome (mtDNA), may impair organism fitness and lead to severe diseases in humans. MtDNA is a multi-copy, highly polymorphic genome that is uniparentally transmitted through the maternal line. Several mechanisms act in the germline to counteract heteroplasmy (i.e., coexistence of two or more mtDNA variants) and prevent expansion of mtDNA mutations. However, reproductive biotechnologies such as cloning by nuclear transfer can disrupt mtDNA inheritance, resulting in new genetic combinations that may be unstable and have physiological consequences. Here, we review the current understanding of mitochondrial inheritance, with emphasis on its pattern in animals and human embryos generated by nuclear transfer.

Keywords:  Cloning; Embryo; Heteroplasmy; MRT; Mitochondria; Nuclear transplantation; Oocyte; SCNT; mtDNA

DOI:  https://doi.org/10.1007/978-1-0716-3064-8_4
iScience. 2023 Apr 21. 26(4): 106386

Tom40 in cholesterol transport.

Himangshu S Bose, Mahuya Bose, Randy M Whittal.

  Cholesterol initiates steroid metabolism in adrenal and gonadal mitochondria, which is essential for all mammalian survival. During stress an increased cholesterol transport rapidly increases steroidogenesis; however, the mechanism of mitochondrial cholesterol transport is unknown. Using rat testicular tissue and mouse Leydig (MA-10) cells, we report for the first time that mitochondrial translocase of outer mitochondrial membrane (OMM), Tom40, is central in cholesterol transport. Cytoplasmic cholesterol-lipids complex containing StAR protein move from the mitochondria-associated ER membrane (MAM) to the OMM, increasing cholesterol load. Tom40 interacts with StAR at the OMM increasing cholesterol transport into mitochondria. An absence of Tom40 disassembles complex formation and inhibits mitochondrial cholesterol transport and steroidogenesis. Therefore, Tom40 is essential for rapid mitochondrial cholesterol transport to initiate, maintain, and regulate activity.

Keywords:  Biomolecules; Cell biology; Protein folding

DOI:  https://doi.org/10.1016/j.isci.2023.106386
Eur J Neurol. 2023 Apr 10.

Phenotyping mtDNA-related diseases in childhood: a cohort study of 150 patients.

Anna Ardissone, Giulia Ferrera, Costanza Lamperti, Valeria Tiranti, Daniele Ghezzi, Isabella Moroni, Eleonora Lamantea.

  BACKGROUND: Mitochondrial diseases (MDs) are heterogeneous disorders caused by mutations in nuclear DNA (nDNA) or mitochondrial DNA (mtDNA) associated with specific syndromes. However, especially in childhood, patients often display heterogeneity. Several reports about the biochemical and molecular profiles in children have been published, but studies tend to not differentiate between mtDNA and nDNA associated diseases and focus is often on a specific phenotype. Thus, large cohort studies specifically focusing on mtDNA defects in the pediatric population are lacking.METHODS: We reviewed the clinical, metabolic, biochemical, and neuroimaging data of 150 patients with MDs due to mtDNA alterations collected at our Neurological Institute over the past 20 years.
RESULTS: MtDNA impairment is less frequent than nDNA in pediatric MDs. Ocular involvement is extremely frequent in our cohort, as is classical Leber Hereditary Optic Neuropathy, especially with onset before 12 years of age. Extra neurological manifestations and isolated myopathy appear to be rare, unlike adult phenotypes. Deep gray matter involvement, early disease onset and specific phenotypes, such as Pearson syndrome and Leigh syndrome, represent unfavorable prognostic factors. Phenotypes related to single large scale mtDNA deletions appear to be very frequent in the pediatric population. Furthermore, we report for the first time a mtDNA pathogenic variant associated with cavitating leukodystrophy.
CONCLUSIONS: We report on a large cohort of pediatric patients with mtDNA defects, adding new data on the phenotypical characterization of mtDNA defects and possible suggestions for the diagnostic workup and therapeutic approach.

Keywords:  mitochondrial DNA; mitochondrial disorder; pediatric; phenotypes

DOI:  https://doi.org/10.1111/ene.15814
bioRxiv. 2023 Mar 31. pii: 2023.03.31.535139. [Epub ahead of print]

An intermembrane space protein facilitates completion of mitochondrial division in yeast.

Olivia M Connor, Srujan K Matta, Jonathan R Friedman.

Mitochondria are highly dynamic double membrane-bound organelles that maintain their shape in part through fission and fusion. Mitochondrial fission is performed by the dynamin-related protein Dnm1 (Drp1 in humans), a large GTPase that constricts and divides the mitochondria in a GTP hydrolysis-dependent manner. However, it is unclear whether factors inside mitochondria help coordinate the process and if Dnm1/Drp1 activity alone is sufficient to complete fission of both mitochondrial membranes. Here, we identify an intermembrane space protein required for mitochondrial fission in yeast, which we propose to name Mdi1. Loss of Mdi1 leads to hyper-fused mitochondria networks due to defects in mitochondrial fission, but not lack of Dnm1 recruitment to mitochondria. Mdi1 plays a conserved role in fungal species and its homologs contain a putative amphipathic α-helix, mutations in which disrupt mitochondrial morphology. One model to explain these findings is that Mdi1 associates with and distorts the mitochondrial inner membrane to enable Dnm1 to robustly complete fission. Our work reveals that Dnm1 cannot efficiently divide mitochondria without the coordinated function of a protein that resides inside mitochondria.

DOI: https://doi.org/10.1101/2023.03.31.535139
J Physiol. 2023 Apr 12.

Increased mitochondrial surface area and cristae density in the skeletal muscle of strength athletes.

Javier Botella, Camilla T Schytz, Thomas F Pehrson, Rune Hokken, Simon Laugesen, Per Aagaard, Charlotte Suetta, Britt Christensen, Niels Ørtenblad, Joachim Nielsen.

  Mitochondria are the cellular organelles responsible for resynthesising the majority of ATP. In skeletal muscle, there is an increased ATP turnover during resistance exercise to sustain the energetic demands of muscle contraction. Despite this, little is known regarding the mitochondrial characteristics of chronically strength-trained individuals and any potential pathways regulating the strength-specific mitochondrial remodelling. Here, we investigated the mitochondrial structural characteristics in skeletal muscle of strength athletes and age-matched untrained controls. The mitochondrial pool in strength athletes was characterised by increased mitochondrial cristae density, decreased mitochondrial size, and increased surface-to-volume ratio, despite similar mitochondrial volume density. We also provide a fibre-type and compartment specific assessment of mitochondria morphology in human skeletal muscle, which reveals across groups a compartment-specific influence on mitochondrial morphology that is largely independent of fibre-type. Furthermore, we show that resistance exercise leads to signs of mild mitochondrial stress, without an increase in the number of damaged mitochondria. Using publicly available transcriptomic data we show that acute resistance exercise increases the expression of markers of mitochondrial biogenesis, fission, and mitochondrial unfolded protein responses (UPRmt ). Further, we observed an enrichment of the UPRmt in the basal transcriptome of strength-trained individuals. Together, these findings show that strength athletes possess a unique mitochondrial remodelling, which minimises the space required for mitochondria. We propose that the concurrent activation of markers of mitochondrial biogenesis and mitochondrial remodelling pathways (fission and UPRmt ) with resistance exercise may be partially responsible for the observed mitochondrial phenotype of strength athletes. KEY POINTS: Untrained individuals and strength athletes possess comparable skeletal muscle mitochondrial volume density. In contrast, strength athletes'mitochondria are characterised by increased cristae density, decreased size, and increased surface-to-volume ratio. Type I fibres have an increased number of mitochondrial profiles with minor differences in the mitochondrial morphological characteristics compared to type II fibres. The mitochondrial morphology is distinct across the subcellular compartments in both groups, with subsarcolemmal mitochondria being bigger in size when compared to intermyofibrillar. Acute resistance exercise leads to signs of mild morphological mitochondrial stress accompanied by increased gene expression of markers of mitochondrial biogenesis, fission, and mitochondrial unfolded protein response (UPRmt). Abstract figure legend: Mitochondria are the cellular organelles responsible for resynthesising the majority of ATP. The present study aimed to investigate the mitochondrial structural characteristics of strength athletes when compared to age-matched untrained individuals. Here we show that the mitochondria of strength athletes have an increased mitochondrial cristae density, increased number of profiles, and an increased surface-to-volume ratio; despite similar mitochondrial volumetric density. Furthermore, we show that human type I fibres, when compared to type II fibres, are characterised by an increased number of mitochondrial profiles without differences in their morphological characteristics. Finally, we show that acute resistance exercise leads to mild signs of mitochondrial morphological stress and an increased expression of markers of mitochondrial biogenesis, fission, and mitochondrial unfolded protein response (UPRmt ). This article is protected by copyright. All rights reserved.

Keywords:  Mitochondria; Olympic weightlifting; cristae density; resistance exercise; skeletal muscle; strength

DOI:  https://doi.org/10.1113/JP284394
Aging (Albany NY). 2023 Apr 10. 15

Mitochondrial redox signaling: a key player in aging and disease.

Maria Vitale, Alberto Sanz, Filippo Scialò.



Keywords:  electron transport chain; mitochondrial reactive oxygen species; mtROS propagation; redox signaling; reverse electron transport

DOI:  https://doi.org/10.18632/aging.204659
bioRxiv. 2023 Mar 29. pii: 2023.03.27.534452. [Epub ahead of print]

A role for N6-methyldeoxyadenosine in C. elegans mitochondrial genome regulation.

Lantana K Grub, James P Held, Tyler J Hansen, Samantha H Schaffner, Marleigh R Canter, Evi M Malagise, Maulik R Patel.

Epigenetic modifications provide powerful means for transmitting information from parent to progeny. As a maternally inherited genome that encodes essential components of the electron transport chain, the mitochondrial genome (mtDNA) is ideally positioned to serve as a conduit for the transgenerational transmission of metabolic information. Here, we provide evidence that mtDNA of C. elegans contains the epigenetic mark N6-methyldeoxyadenosine (6mA). Bioinformatic analysis of SMRT sequencing data and methylated DNA IP sequencing data reveal that C. elegans mtDNA is methylated at high levels in a site-specific manner. We further confirmed that mtDNA contains 6mA by leveraging highly specific anti-6mA antibodies. Additionally, we find that mtDNA methylation is dynamically regulated in response to antimycin, a mitochondrial stressor. Further, 6mA is increased in nmad-1 mutants and is accompanied by a significant decrease in mtDNA copy number. Our discovery paves the way for future studies to investigate the regulation and inheritance of mitochondrial epigenetics.

DOI: https://doi.org/10.1101/2023.03.27.534452
Cells. 2023 Apr 05. pii: 1089. [Epub ahead of print]12(7):

High-Throughput Microscopy Analysis of Mitochondrial Membrane Potential in 2D and 3D Models.

Caterina Vianello, Federica Dal Bello, Sang Hun Shin, Sara Schiavon, Camilla Bean, Ana Paula Magalhães Rebelo, Tomáš Knedlík, Emad Norouzi Esfahani, Veronica Costiniti, Rodrigo S Lacruz, Giuseppina Covello, Fabio Munari, Tommaso Scolaro, Antonella Viola, Elena Rampazzo, Luca Persano, Sara Zumerle, Luca Scorrano, Alessio Gianelle, Marta Giacomello.

  Recent proteomic, metabolomic, and transcriptomic studies have highlighted a connection between changes in mitochondria physiology and cellular pathophysiological mechanisms. Secondary assays to assess the function of these organelles appear fundamental to validate these -omics findings. Although mitochondrial membrane potential is widely recognized as an indicator of mitochondrial activity, high-content imaging-based approaches coupled to multiparametric to measure it have not been established yet. In this paper, we describe a methodology for the unbiased high-throughput quantification of mitochondrial membrane potential in vitro, which is suitable for 2D to 3D models. We successfully used our method to analyze mitochondrial membrane potential in monolayers of human fibroblasts, neural stem cells, spheroids, and isolated muscle fibers. Moreover, by combining automated image analysis and machine learning, we were able to discriminate melanoma cells from macrophages in co-culture and to analyze the subpopulations separately. Our data demonstrated that our method is a widely applicable strategy for large-scale profiling of mitochondrial activity.

Keywords:  NSCs; TMRM; co-culture; machine learning; mitochondrial membrane potential; single muscle fibers; spheroids

DOI:  https://doi.org/10.3390/cells12071089
J Mol Biol. 2023 Apr 11. pii: S0022-2836(23)00152-3. [Epub ahead of print] 168090

Structural mechanisms of mitochondrial quality control mediated by PINK1 and parkin.

Jean-François Trempe, Kalle Gehring.

Parkinson's disease (PD) is the second most common neurodegenerative disease and represents a looming public health crisis as the global population ages. While the etiology of the more common, idiopathic form of the disease remains unknown, the last ten years have seen a breakthrough in our understanding of the genetic forms related to two proteins that regulate a quality control system for the removal of damaged or non-functional mitochondria. Here, we review the structure of these proteins, PINK1, a protein kinase, and parkin, a ubiquitin ligase with an emphasis on the molecular mechanisms responsible for their recognition of dysfunctional mitochondria and control of the subsequent ubiquitination cascade. Recent atomic structures have revealed the basis of PINK1 substrate specificity and the conformational changes responsible for activation of PINK1 and parkin catalytic activity. Progress in understanding the molecular basis of mitochondrial quality control promises to open new avenues for therapeutic interventions in PD.

DOI: https://doi.org/10.1016/j.jmb.2023.168090
Genet Med. 2023 Apr 10. pii: S1098-3600(23)00851-1. [Epub ahead of print] 100838

De novo missense variants in RRAGC lead to a fatal mTORopathy of early childhood.

Margot R F Reijnders, Annette Seibt, Melanie Brugger, Ideke J C Lamers, Torsten Ott, Oliver Klaas, Judit Horváth, Ailsa M S Rose, Isabel M Craghill, Theresa Brunet, Elisabeth Graf, Katharina Mayerhanser, Debby Hellebrekers, David Pauck, Eva Neuen-Jacob, Richard J T Rodenburg, Dagmar Wieczorek, Dirk Klee, Ertan Mayatepek, Gertjan Driessen, Robert Bindermann, Luisa Averdunk, Klaus Lohmeier, Margje Sinnema, Alexander P A Stegmann, Ronald Roepman, James A Poulter, Felix Distelmaier.

  INTRODUCTION: Mechanistic target of rapamycin complex 1 (mTORC1) regulates cell growth in response to nutritional status. Central to mTORC1 function is the Rag-GTPase heterodimer. One component of the Rag heterodimer is RagC (Ras-related GTP-binding protein C), which is encoded by the RRAGC gene.MATERIAL AND METHODS: Genetic testing via trio exome sequencing was applied to identify the underlying disease cause in three infants who suffered from dilated cardiomyopathy, hepatopathy and brain abnormalities including pachygyria, polymicrogyria, and septo-optic dysplasia. Studies in patient-derived skin fibroblasts and in a HEK293 cell model were performed to investigate the cellular consequences.
RESULTS: We identified three de novo missense variants in RRAGC (NM_022157.4: c.269C>A, p.(Thr90Asn), c.353C>T, p.(Pro118Leu), and c.343T>C, p.(Trp115Arg)), which were previously reported as occurring somatically in follicular lymphoma. Studies of patient-derived fibroblasts carrying the p.(Thr90Asn) variant revealed increased cell size as well as dysregulation of mTOR-related p70S6K (ribosomal protein S6 kinase 1) and TFEB (transcription factor EB) signaling. Moreover, subcellular localization of mTOR was decoupled from metabolic state. We confirmed the key-findings for all RRAGC variants described in this study in a HEK293 cell model.
DISCUSSION: The above results are in line with a constitutive over-activation of the mTORC1 pathway. Our study establishes de novo missense variants in RRAGC as cause of an early-onset mTORopathy with unfavorable prognosis.

Keywords:  cardiomyopathy; cortical malformation; heart; lysosome; mTORopathy; mitochondrial

DOI:  https://doi.org/10.1016/j.gim.2023.100838
J Med Genet. 2023 Apr 13. pii: jmg-2022-109027. [Epub ahead of print]

Strategic validation of variants of uncertain significance in ECHS1 genetic testing.

Yoshihito Kishita, Ayumu Sugiura, Takanori Onuki, Tomohiro Ebihara, Tetsuro Matsuhashi, Masaru Shimura, Takuya Fushimi, Noriko Ichino, Yoshie Nagatakidani, Hitomi Nishihata, Kazuhiro R Nitta, Yukiko Yatsuka, Atsuko Imai-Okazaki, Yibo Wu, Hitoshi Osaka, Akira Ohtake, Kei Murayama, Yasushi Okazaki.

  BACKGROUND: Enoyl-CoA hydratase short-chain 1 (ECHS1) is an enzyme involved in the metabolism of branched chain amino acids and fatty acids. Mutations in the ECHS1 gene lead to mitochondrial short-chain enoyl-CoA hydratase 1 deficiency, resulting in the accumulation of intermediates of valine. This is one of the most common causative genes in mitochondrial diseases. While genetic analysis studies have diagnosed numerous cases with ECHS1 variants, the increasing number of variants of uncertain significance (VUS) in genetic diagnosis is a major problem.METHODS: Here, we constructed an assay system to verify VUS function for ECHS1 gene. A high-throughput assay using ECHS1 knockout cells was performed to index these phenotypes by expressing cDNAs containing VUS. In parallel with the VUS validation system, a genetic analysis of samples from patients with mitochondrial disease was performed. The effect on gene expression in cases was verified by RNA-seq and proteome analysis.
RESULTS: The functional validation of VUS identified novel variants causing loss of ECHS1 function. The VUS validation system also revealed the effect of the VUS in the compound heterozygous state and provided a new methodology for variant interpretation. Moreover, we performed multiomics analysis and identified a synonymous substitution p.P163= that results in splicing abnormality. The multiomics analysis complemented the diagnosis of some cases that could not be diagnosed by the VUS validation system.
CONCLUSIONS: In summary, this study uncovered new ECHS1 cases based on VUS validation and omics analysis; these analyses are applicable to the functional evaluation of other genes associated with mitochondrial disease.

Keywords:  RNA-Seq; genetic testing; genetic variation; molecular diagnostic techniques

DOI:  https://doi.org/10.1136/jmg-2022-109027
Nat Rev Genet. 2023 Apr 14.

Variant calling and benchmarking in an era of complete human genome sequences.

Nathan D Olson, Justin Wagner, Nathan Dwarshuis, Karen H Miga, Fritz J Sedlazeck, Marc Salit, Justin M Zook.

Genetic variant calling from DNA sequencing has enabled understanding of germline variation in hundreds of thousands of humans. Sequencing technologies and variant-calling methods have advanced rapidly, routinely providing reliable variant calls in most of the human genome. We describe how advances in long reads, deep learning, de novo assembly and pangenomes have expanded access to variant calls in increasingly challenging, repetitive genomic regions, including medically relevant regions, and how new benchmark sets and benchmarking methods illuminate their strengths and limitations. Finally, we explore the possible future of more complete characterization of human genome variation in light of the recent completion of a telomere-to-telomere human genome reference assembly and human pangenomes, and we consider the innovations needed to benchmark their newly accessible repetitive regions and complex variants.

DOI: https://doi.org/10.1038/s41576-023-00590-0
Front Neurosci. 2023 ;17 1119724

Comparison of different gene-therapy methods to treat Leber hereditary optic neuropathy in a mouse model.

Sindhu Velmurugan, Tsung-Han Chou, Jeremy D Eastwood, Vittorio Porciatti, Yuan Liu, William W Hauswirth, John Guy, Hong Yu.

  Introduction: Therapies for Leber hereditary optic neuropathy (LHON), in common with all disorders caused by mutated mitochondrial DNA, are inadequate. We have developed two gene therapy strategies for the disease: mitochondrial-targeted and allotopic expressed and compared them in a mouse model of LHON.Methods: A LHON mouse model was generated by intravitreal injection of a mitochondrialtargeted Adeno-associated virus (AAV) carrying mutant human NADH dehydrogenase 4 gene (hND4/m.11778G>A) to induce retinal ganglion cell (RGC) degeneration and axon loss, the hallmark of the human disease. We then attempted to rescue those mice using a second intravitreal injection of either mitochondrial-targeted or allotopic expressed wildtype human ND4. The rescue of RGCs and their axons were assessed using serial pattern electroretinogram (PERG) and transmission electron microscopy.
Results: Compared to non-rescued LHON controls where PERG amplitude was much reduced, both strategies significantly preserved PERG amplitude over 15 months. However, the rescue effect was more marked with mitochondrial-targeted therapy than with allotopic therapy (p = 0.0128). Post-mortem analysis showed that mitochondrial-targeted human ND4 better preserved small axons that are preferentially lost in human LHON.
Conclusions: These results in a pre-clinical mouse model of LHON suggest that mitochondrially-targeted AAV gene therapy, compared to allotopic AAV gene therapy, is more efficient in rescuing the LHON phenotype.

Keywords:  Leber hereditary optic neuropathy (LHON); allotopic expression; gene therapy; mitochondrial-targeted; mitochondrial-targeted therapy

DOI:  https://doi.org/10.3389/fnins.2023.1119724
Front Cell Dev Biol. 2023 ;11 1107930

A retrospective analysis of phosphatase catalytic subunit gene variants in patients with rare disorders identifies novel candidate neurodevelopmental disease genes.

Ekaterina Lyulcheva-Bennett, Genomics England Research Consortium, Daimark Bennett.

  Rare genetic disorders represent some of the most severe and life-limiting conditions that constitute a considerable burden on global healthcare systems and societies. Most individuals affected by rare disorders remain undiagnosed, highlighting the unmet need for improved disease gene discovery and novel variant interpretation. Aberrant (de) phosphorylation can have profound pathological consequences underpinning many disease processes. Numerous phosphatases and associated proteins have been identified as disease genes, with many more likely to have gone undiscovered thus far. To begin to address these issues, we have performed a systematic survey of de novo variants amongst 189 genes encoding phosphatase catalytic subunits found in rare disease patients recruited to the 100,000 Genomes Project (100 kGP), the largest national sequencing project of its kind in the United Kingdom. We found that 49% of phosphatases were found to carry de novo mutation(s) in this cohort. Only 25% of these phosphatases have been previously linked to genetic disorders. A gene-to-patient approach matching variants to phenotypic data identified 9 novel candidate rare-disease genes: PTPRD, PTPRG, PTPRT, PTPRU, PTPRZ1, MTMR3, GAK, TPTE2, PTPN18. As the number of patients undergoing whole genome sequencing increases and information sharing improves, we anticipate that reiterative analysis of genomic and phenotypic data will continue to identify candidate phosphatase disease genes for functional validation. This is the first step towards delineating the aetiology of rare genetic disorders associated with altered phosphatase function, leading to new biological insights and improved clinical outcomes for the affected individuals and their families.

Keywords:  de novo mutation; developmental disorder; disease genes; genome sequencing; mendelian disease; phosphatase; phosphatome; rare disorders

DOI:  https://doi.org/10.3389/fcell.2023.1107930
Cell Calcium. 2023 Apr 05. pii: S0143-4160(23)00048-9. [Epub ahead of print]112 102736

Synergy of uncoupling proteins (1 and 2) with mitochondrial Ca2+ uptake machinery potentiate mitochondrial uncoupling.

Furkan E Oflaz, Zhanat Koshenov, Martin Hirtl, Olaf A Bachkoenig, Wolfgang F Graier, Benjamin Gottschalk.

  Mitochondrial uncoupling proteins UCP1 and UCP2 have a structural homology of app. 60%. They execute their mitochondria uncoupling function through different molecular mechanisms. Non-shivering thermogenesis by UCP1 is mediated through a transmembrane dissipation of the proton motive force to create heat during sympathetic stimulation. UCP2, on the other hand, modulates through the interaction with methylated MICU1 the permeability of the cristae junction, which acts as an isolator for the cristae-located mitochondrial membrane potential. In this mini-review, we discuss and compare the recently described molecular mechanism of UCP1 in brown adipose tissue and UCP2 in aged and cancer non-excitable cells that contribute to mitochondrial uncoupling, and the synergistic effects of both UCPs with the mitochondrial Ca2+ uptake machinery.

Keywords:  Mitochondrial Ca(2+) uptake machinery; Mitochondrial uncoupling; Uncoupling protein 1; Uncoupling protein 2

DOI:  https://doi.org/10.1016/j.ceca.2023.102736
Int J Mol Sci. 2023 Apr 01. pii: 6593. [Epub ahead of print]24(7):

PI4P-Containing Vesicles from Golgi Contribute to Mitochondrial Division by Coordinating with Polymerized Actin.

Xinxin Duan, Yunfei Wei, Meng Zhang, Wenting Zhang, Yue Huang, Yu-Hui Zhang.

  Golgi-derived PI4P-containing vesicles play important roles in mitochondrial division, which is essential for maintaining cellular homeostasis. However, the mechanism of the PI4P-containing vesicle effect on mitochondrial division is unclear. Here, we found that actin appeared to polymerize at the contact site between PI4P-containing vesicles and mitochondria, causing mitochondrial division. Increasing the content of PI4P derived from the Golgi apparatus increased actin polymerization and reduced the length of the mitochondria, suggesting that actin polymerization through PI4P-containing vesicles is involved in PI4P vesicle-related mitochondrial division. Collectively, our results support a model in which PI4P-containing vesicles derived from the Golgi apparatus cooperate with actin filaments to participate in mitochondrial division by contributing to actin polymerization, which regulates mitochondrial dynamics. This study enriches the understanding of the pathways that regulate mitochondrial division and provides new insight into mitochondrial dynamics.

Keywords:  PI4P-containing vesicle; actin polymerization; interaction; mitochondrial division; super-resolution imaging

DOI:  https://doi.org/10.3390/ijms24076593
Eur Heart J. 2023 Apr 13. pii: ehad177. [Epub ahead of print]

Mitochondrial cardiomyopathy: a fertile field for research.

Eugene Braunwald.

DOI: https://doi.org/10.1093/eurheartj/ehad177
Nat Cell Biol. 2023 Apr;25(4): 592-603

Arg-tRNA synthetase links inflammatory metabolism to RNA splicing and nuclear trafficking via SRRM2.

Haissi Cui, Jolene K Diedrich, Douglas C Wu, Justin J Lim, Ryan M Nottingham, James J Moresco, John R Yates, Benjamin J Blencowe, Alan M Lambowitz, Paul Schimmel.

Cells respond to perturbations such as inflammation by sensing changes in metabolite levels. Especially prominent is arginine, which has known connections to the inflammatory response. Aminoacyl-tRNA synthetases, enzymes that catalyse the first step of protein synthesis, can also mediate cell signalling. Here we show that depletion of arginine during inflammation decreased levels of nuclear-localized arginyl-tRNA synthetase (ArgRS). Surprisingly, we found that nuclear ArgRS interacts and co-localizes with serine/arginine repetitive matrix protein 2 (SRRM2), a spliceosomal and nuclear speckle protein, and that decreased levels of nuclear ArgRS correlated with changes in condensate-like nuclear trafficking of SRRM2 and splice-site usage in certain genes. These splice-site usage changes cumulated in the synthesis of different protein isoforms that altered cellular metabolism and peptide presentation to immune cells. Our findings uncover a mechanism whereby an aminoacyl-tRNA synthetase cognate to a key amino acid that is metabolically controlled during inflammation modulates the splicing machinery.

DOI: https://doi.org/10.1038/s41556-023-01118-8
Cells. 2023 Mar 23. pii: 985. [Epub ahead of print]12(7):

Tau Transfer via Extracellular Vesicles Disturbs the Astrocytic Mitochondrial System.

Romain Perbet, Valentin Zufferey, Elodie Leroux, Enea Parietti, Jeanne Espourteille, Lucas Culebras, Sylvain Perriot, Renaud Du Pasquier, Séverine Bégard, Vincent Deramecourt, Nicole Déglon, Nicolas Toni, Luc Buée, Morvane Colin, Kevin Richetin.

  Tauopathies are neurodegenerative disorders involving the accumulation of tau isoforms in cell subpopulations such as astrocytes. The origins of the 3R and 4R isoforms of tau that accumulate in astrocytes remain unclear. Extracellular vesicles (EVs) were isolated from primary neurons overexpressing 1N3R or 1N4R tau or from human brain extracts (progressive supranuclear palsy or Pick disease patients or controls) and characterized (electron microscopy, nanoparticle tracking analysis (NTA), proteomics). After the isolated EVs were added to primary astrocytes or human iPSC-derived astrocytes, tau transfer and mitochondrial system function were evaluated (ELISA, immunofluorescence, MitoTracker staining). We demonstrated that neurons in which 3R or 4R tau accumulated had the capacity to transfer tau to astrocytes and that EVs were essential for the propagation of both isoforms of tau. Treatment with tau-containing EVs disrupted the astrocytic mitochondrial system, altering mitochondrial morphology, dynamics, and redox state. Although similar levels of 3R and 4R tau were transferred, 3R tau-containing EVs were significantly more damaging to astrocytes than 4R tau-containing EVs. Moreover, EVs isolated from the brain fluid of patients with different tauopathies affected mitochondrial function in astrocytes derived from human iPSCs. Our data indicate that tau pathology spreads to surrounding astrocytes via EVs-mediated transfer and modifies their function.

Keywords:  astrocytes; extracellular vesicles; mitochondria; tau spreading; tauopathies

DOI:  https://doi.org/10.3390/cells12070985
Nat Cell Biol. 2023 Apr 10.

Genome-wide CRISPR screens identify ILF3 as a mediator of mTORC1-dependent amino acid sensing.

Guokai Yan, Jinxin Yang, Wen Li, Ao Guo, Jialiang Guan, Ying Liu.

The mechanistic target of rapamycin complex 1 (mTORC1) is an essential hub that integrates nutrient signals and coordinates metabolism to control cell growth. Amino acid signals are detected by sensor proteins and relayed to the GATOR2 and GATOR1 complexes to control mTORC1 activity. Here we perform genome-wide CRISPR/Cas9 screens, coupled with an assay for mTORC1 activity based on fluorescence-activated cell sorting analysis of pS6, to identify potential regulators of mTORC1-dependent amino acid sensing. We then focus on interleukin enhancer binding factor 3 (ILF3), one of the candidate genes from the screen. ILF3 tethers the GATOR complexes to lysosomes to control mTORC1. Adding a lysosome-targeting sequence to the GATOR2 component WDR24 bypasses the requirement for ILF3 to modulate amino-acid-dependent mTORC1 signalling. ILF3 plays an evolutionarily conserved role in human and mouse cells, and in worms to regulate the mTORC1 pathway, control autophagy activity and modulate the ageing process.

DOI: https://doi.org/10.1038/s41556-023-01123-x
Curr Protoc. 2023 Apr;3(4): e720

Clinical Untargeted Metabolomics as a Functional Screen to Improve Variant Classification.

Charul Gijavanekar, Sarah H Elsea.

  With the rapid increase in clinical exome and genome sequencing, the number of variants of uncertain significance (VUS) that are reported continues to rise, which poses a significant barrier to interpretation of genetic findings. For metabolic disorders, biochemical testing can help alleviate this burden of variant interpretation by providing functional validation of uncertain genetic findings in many cases. However, a major limitation of traditional biochemical testing is the targeted, narrow range of analytes clinically available, resulting in delays in diagnosis if testing is negative. Untargeted metabolomic screening offers higher diagnostic yield and assays for thousands of metabolites across multiple metabolic pathways in a single test, saving time and resources for patients, families, and physicians. When integrated with exome or genome sequencing, untargeted metabolomic screening improves diagnostic outcomes by providing functional validation of genetic findings, particularly for VUS. Here, we present representative cases across the breadth of metabolic pathways as examples of the utility of metabolomics in genomic variant classification. © 2023 Wiley Periodicals LLC.

Keywords:  biochemical genetics; functional screen; functional validation; untargeted metabolomics; variant classification; variants of uncertain significance

DOI:  https://doi.org/10.1002/cpz1.720
Wiley Interdiscip Rev RNA. 2023 Apr 12. e1789

Physiological and engineered tRNA aminoacylation.

Santiago Tijaro-Bulla, Samuel Protais Nyandwi, Haissi Cui.

  Aminoacyl-tRNA synthetases form the protein family that controls the interpretation of the genetic code, with tRNA aminoacylation being the key chemical step during which an amino acid is assigned to a corresponding sequence of nucleic acids. In consequence, aminoacyl-tRNA synthetases have been studied in their physiological context, in disease states, and as tools for synthetic biology to enable the expansion of the genetic code. Here, we review the fundamentals of aminoacyl-tRNA synthetase biology and classification, with a focus on mammalian cytoplasmic enzymes. We compile evidence that the localization of aminoacyl-tRNA synthetases can be critical in health and disease. In addition, we discuss evidence from synthetic biology which made use of the importance of subcellular localization for efficient manipulation of the protein synthesis machinery. This article is categorized under: RNA Processing Translation > Translation Regulation RNA Processing > tRNA Processing RNA Export and Localization > RNA Localization.

Keywords:  genetic code expansion; mRNA translation; neurological disorders; synthetic biology; tRNA

DOI:  https://doi.org/10.1002/wrna.1789
Sci Adv. 2023 Apr 14. 9(15): eadf1956

Defective import of mitochondrial metabolic enzyme elicits ectopic metabolic stress.

Kazuya Nishio, Tomoyuki Kawarasaki, Yuki Sugiura, Shunsuke Matsumoto, Ayano Konoshima, Yuki Takano, Mayuko Hayashi, Fumihiko Okumura, Takumi Kamura, Tsunehiro Mizushima, Kunio Nakatsukasa.

Deficiencies in mitochondrial protein import are associated with a number of diseases. However, although nonimported mitochondrial proteins are at great risk of aggregation, it remains largely unclear how their accumulation causes cell dysfunction. Here, we show that nonimported citrate synthase is targeted for proteasomal degradation by the ubiquitin ligase SCFUcc1. Unexpectedly, our structural and genetic analyses revealed that nonimported citrate synthase appears to form an enzymatically active conformation in the cytosol. Its excess accumulation caused ectopic citrate synthesis, which, in turn, led to an imbalance in carbon flux of sugar, a reduction of the pool of amino acids and nucleotides, and a growth defect. Under these conditions, translation repression is induced and acts as a protective mechanism that mitigates the growth defect. We propose that the consequence of mitochondrial import failure is not limited to proteotoxic insults, but that the accumulation of a nonimported metabolic enzyme elicits ectopic metabolic stress.

DOI: https://doi.org/10.1126/sciadv.adf1956
Proc Natl Acad Sci U S A. 2023 Apr 18. 120(16): e2217665120

Evidence supporting the MICU1 occlusion mechanism and against the potentiation model in the mitochondrial calcium uniporter complex.

Chen-Wei Tsai, Tsung-Yun Liu, Fan-Yi Chao, Yung-Chi Tu, Madison X Rodriguez, Anna M Van Keuren, Zhiwei Ma, John Bankston, Ming-Feng Tsai.

  The mitochondrial calcium uniporter is a Ca2+ channel that imports cytoplasmic Ca2+ into the mitochondrial matrix to regulate cell bioenergetics, intracellular Ca2+ signaling, and apoptosis. The uniporter contains the pore-forming MCU subunit, an auxiliary EMRE protein, and the regulatory MICU1/MICU2 subunits. Structural and biochemical studies have suggested that MICU1 gates MCU by blocking/unblocking the pore. However, mitoplast patch-clamp experiments argue that MICU1 does not block, but instead potentiates MCU via allosteric mechanisms. Here, we address this direct clash of the proposed MICU1 function. Supporting the MICU1-occlusion mechanism, patch-clamp demonstrates that purified MICU1 strongly suppresses MCU Ca2+ currents, and this inhibition is abolished by mutating the MCU-interacting K126 residue. Moreover, a membrane-depolarization assay shows that MICU1 prevents MCU-mediated Na+ flux into intact mitochondria under Ca2+-free conditions. Examining the observations underlying the potentiation model, we found that MICU1 occlusion was not detected in mitoplasts not because MICU1 cannot block, but because MICU1 dissociates from the uniporter complex. Furthermore, MICU1 depletion reduces uniporter transport not because MICU1 can potentiate MCU, but because EMRE is down-regulated. These results firmly establish the molecular mechanisms underlying the physiologically crucial process of uniporter regulation by MICU1.

Keywords:  calcium channels; intracellular calcium signaling; mitochondrial physiology; organellar channels

DOI:  https://doi.org/10.1073/pnas.2217665120
Nat Commun. 2023 Apr 11. 14(1): 2030

Cryo-EM structures of mitochondrial ABC transporter ABCB10 in apo and biliverdin-bound form.

Sheng Cao, Yihu Yang, Lili He, Yumo Hang, Xiaodong Yan, Hui Shi, Jiaquan Wu, Zhuqing Ouyang.

ABCB10, a member of ABC transporter superfamily that locates in the inner membrane of mitochondria, plays crucial roles in hemoglobin synthesis, antioxidative stress and stabilization of the iron transporter mitoferrin-1. Recently, it was found that ABCB10 is a mitochondrial biliverdin exporter. However, the molecular mechanism of biliverdin export by ABCB10 remains elusive. Here we report the cryo-EM structures of ABCB10 in apo (ABCB10-apo) and biliverdin-bound form (ABCB10-BV) at 3.67 Å and 2.85 Å resolution, respectively. ABCB10-apo adopts a wide-open conformation and may thus represent the apo form structure. ABCB10-BV forms a closed conformation and biliverdin situates in a hydrophobic pocket in one protomer and bridges the interaction through hydrogen bonds with the opposing one. We also identify cholesterols sandwiched by BVs and discuss the export dynamics based on these structural and biochemical observations.

DOI: https://doi.org/10.1038/s41467-023-37851-9
Front Endocrinol (Lausanne). 2023 ;14 1122012

Mitochondrial stress response gene Clpp deficiency impairs oocyte competence and deteriorate cyclophosphamide-induced ovarian damage in young mice.

Guangxin Li, Jingkai Gu, Xiaomei Zhou, Ting Wu, Xian Li, Renwu Hua, Zhuo Hai, Yuan Xiao, Jiaping Su, Willian S B Yeung, Kui Liu, Chenxi Guo, Tianren Wang.

  Chemotherapy is extensively used to treat cancers and is often associated with ovarian damage and leads to premature ovarian insufficiency and infertility, while the role of mitochondria during ovarian damage with chemotherapy remains unknown. This study used a mouse model with oocyte-specific deletion of mitochondrial stress response gene Caseinolytic peptidase P (Clpp) to investigate mitochondrial homeostasis in oocytes from mice receiving a chemotherapeutic drug cyclophosphamide (CTX). We found that oocyte-specific deletion of Clpp reduced fecundity of the mice at advanced age. The deletion led to meiotic defects with elevated abnormal spindle rate and aneuploidy rate with impaired mitochondrial function in the MII oocytes from 8-week-old mice. Upon CTX treatment at 8-week-old, the oocyte competence and folliculogenesis from the oocyte-specific Clpp knockout mice was further deteriorated with dramatic impairment of mitochondrial distribution and function including elevated ROS level, decreased mitochondrial membrane potential, respiratory chain activity and ATP production. Taken together, the results indicate that that ClpP was required for oocyte competence during maturation and early folliculogenesis, and its deficiency deteriorate cyclophosphamide-induced ovarian damage.

Keywords:  ClpP; cyclophosphamide; follicle development; mitochondria; oocyte

DOI:  https://doi.org/10.3389/fendo.2023.1122012
Neurol Sci. 2023 Apr 12.

FDXR-associated disease: a challenging differential diagnosis with inflammatory peripheral neuropathy.

Silvia Masnada, Roberto Previtali, Paola Erba, Elena Beretta, Anna Camporesi, Luisa Chiapparini, Chiara Doneda, Maria Iascone, Marco U A Sartorio, Luigina Spaccini, Pierangelo Veggiotti, Maurizio Osio, Davide Tonduti, Isabella Moroni.

  BACKGROUND AND AIMS: Mutations in FDXR gene, involved in mitochondrial pathway, cause a rare recessive neurological disorder with variable severity of phenotypes. The most common presentation includes optic and/or auditory neuropathy, variably associated to developmental delay or regression, global hypotonia, pyramidal, cerebellar signs, and seizures. The review of clinical findings in previously described cases from literature reveals also a significant incidence of sensorimotor peripheral polyneuropathy (22.72%) and ataxia (43.18%). To date, 44 patients with FDXR mutations have been reported. We describe here on two new patients, siblings, who presented with a quite different phenotype compared to previously described patients.METHODS: Clinical, neurophysiological, and genetic features of two siblings and a systematic literature review focused on the clinical spectrum of the disease are described.
RESULTS: Both patients presented with an acute-sub-acute onset of peripheral neuropathy and only in later stages of the disease developed the typical features of FDXR-associated disease.
INTERPRETATION: The peculiar clinical presentation at onset and the evolution of the disease in our patients and in some cases revised from the literature shed lights on a new possible phenotype of FDXR-associated disease: a peripheral neuropathy which can mimic an acute inflammatory disease.

Keywords:  FDXR; Peripheral neuropathy; Phenotype

DOI:  https://doi.org/10.1007/s10072-023-06790-0
medRxiv. 2023 Mar 30. pii: 2023.03.21.23287545. [Epub ahead of print]

A gene pathogenicity tool 'GenePy' identifies missed biallelic diagnoses in the 100,000 Genomes Project.

Genomics England Consortium

The 100,000 Genomes Project (100KGP) diagnosed a quarter of recruited affected participants, but 26% of diagnoses were in genes not on the chosen gene panel(s); with many being de novo variants of high impact. However, assessing biallelic variants without a gene panel is challenging, due to the number of variants requiring scrutiny. We sought to identify potential missed biallelic diagnoses independent of the gene panel applied using GenePy - a whole gene pathogenicity metric. GenePy scores all variants called in a given individual, incorporating allele frequency, zygosity, and a user-defined deleterious metric (CADD v1.6 applied herein). GenePy then combines all variant scores for individual genes, generating an aggregate score per gene, per participant. We calculated GenePy scores for 2862 recessive disease genes in 78,216 individuals in 100KGP. For each gene, we ranked participant GenePy scores for that gene, and scrutinised affected individuals without a diagnosis whose scores ranked amongst the top-5 for each gene. We assessed these participants' phenotypes for overlap with the disease gene associated phenotype for which they were highly ranked. Where phenotypes overlapped, we extracted rare variants in the gene of interest and applied phase, ClinVar and ACMG classification looking for putative causal biallelic variants. 3184 affected individuals without a molecular diagnosis had a top-5 ranked GenePy gene score and 682/3184 (21%) had phenotypes overlapping with one of the top-ranking genes. After removing 13 withdrawn participants, in 122/669 (18%) of the phenotype-matched cases, we identified a putative missed diagnosis in a top-ranked gene supported by phasing, ClinVar and ACMG classification. A further 334/669 (50%) of cases have a possible missed diagnosis but require functional validation. Applying GenePy at scale has identified potential diagnoses for 456/3183 (14%) of undiagnosed participants who had a top-5 ranked GenePy score in a recessive disease gene, whilst adding only 1.2 additional variants (per individual) for assessment.

DOI: https://doi.org/10.1101/2023.03.21.23287545
Arch Clin Cases. 2023 ;10(1): 42-46

From recurrent rhabdomyolysis in a young adult to carnitine palmitoyltransferase II deficiency.

Cristina Marques, Catarina Silva, Carina Silva, João Pedro Abreu, Márcia Ribeiro, Arlindo Guimas.

  Metabolic myopathies are a diverse group of rare genetic disorders associated with recurrent episodes of rhabdomyolysis, induced by triggers such as fever or exercise. In these disorders, the energetic metabolism is compromised resulting in damage of the muscle cells. The diagnosis can be challenging but is essential for the correct treatment. Carnitine palmitoytransferase II (CPT-II) deficiency is the most common long-chain fatty acid oxidation defect, with the adulthood form requiring additional external triggers. The authors present a case of a young-male adult with recurrent episodes of rhabdomyolysis, one of them presented with acute renal failure and acute hepatitis. The diagnostic is demanding, which requires a high level of suspicion. The adequate treatment of these patients improves the muscle function and prevents other episodes of severe rhabdomyolysis.

Keywords:  CPT II deficiency; acute renal failure; hepatitis; rhabdomyolysis

DOI:  https://doi.org/10.22551/2023.38.1001.10238
Int J Mol Sci. 2023 Mar 24. pii: 6128. [Epub ahead of print]24(7):

Mitochondrial Signaling Pathways Associated with DNA Damage Responses.

Tsutomu Shimura.

  Under physiological and stress conditions, mitochondria act as a signaling platform to initiate biological events, establishing communication from the mitochondria to the rest of the cell. Mitochondrial adenosine triphosphate (ATP), reactive oxygen species, cytochrome C, and damage-associated molecular patterns act as messengers in metabolism, oxidative stress response, bystander response, apoptosis, cellular senescence, and inflammation response. In this review paper, the mitochondrial signaling in response to DNA damage was summarized. Mitochondrial clearance via fusion, fission, and mitophagy regulates mitochondrial quality control under oxidative stress conditions. On the other hand, damaged mitochondria release their contents into the cytoplasm and then mediate various signaling pathways. The role of mitochondrial dysfunction in radiation carcinogenesis was discussed, and the recent findings on radiation-induced mitochondrial signaling and radioprotective agents that targeted mitochondria were presented. The analysis of the mitochondrial radiation effect, as hypothesized, is critical in assessing radiation risks to human health.

Keywords:  DNA damage response; inflammation response; mitochondrial signaling; oxidative stress; radiation carcinogenesis

DOI:  https://doi.org/10.3390/ijms24076128
Nat Commun. 2023 Apr 14. 14(1): 2123

IDH3γ functions as a redox switch regulating mitochondrial energy metabolism and contractility in the heart.

Maithily S Nanadikar, Ana M Vergel Leon, Jia Guo, Gijsbert J van Belle, Aline Jatho, Elvina S Philip, Astrid F Brandner, Rainer A Böckmann, Runzhu Shi, Anke Zieseniss, Carla M Siemssen, Katja Dettmer, Susanne Brodesser, Marlen Schmidtendorf, Jingyun Lee, Hanzhi Wu, Cristina M Furdui, Sören Brandenburg, Joseph R Burgoyne, Ivan Bogeski, Jan Riemer, Arpita Chowdhury, Peter Rehling, Tobias Bruegmann, Vsevolod V Belousov, Dörthe M Katschinski.

Redox signaling and cardiac function are tightly linked. However, it is largely unknown which protein targets are affected by hydrogen peroxide (H2O2) in cardiomyocytes that underly impaired inotropic effects during oxidative stress. Here, we combine a chemogenetic mouse model (HyPer-DAO mice) and a redox-proteomics approach to identify redox sensitive proteins. Using the HyPer-DAO mice, we demonstrate that increased endogenous production of H2O2 in cardiomyocytes leads to a reversible impairment of cardiac contractility in vivo. Notably, we identify the γ-subunit of the TCA cycle enzyme isocitrate dehydrogenase (IDH)3 as a redox switch, linking its modification to altered mitochondrial metabolism. Using microsecond molecular dynamics simulations and experiments using cysteine-gene-edited cells reveal that IDH3γ Cys148 and 284 are critically involved in the H2O2-dependent regulation of IDH3 activity. Our findings provide an unexpected mechanism by which mitochondrial metabolism can be modulated through redox signaling processes.

DOI: https://doi.org/10.1038/s41467-023-37744-x
Circ Res. 2023 Apr 14.

Enhanced Mitochondria-SR Tethering Triggers Adaptive Cardiac Muscle Remodeling.

Zuzana Nichtová, Celia Fernandez-Sanz, Sergio De La Fuente, Yuexing Yuan, Stephen Hurst, Sebastian Lanvermann, Hui-Ying Tsai, David Weaver, Ariele Baggett, Christopher Thompson, Cedric Bouchet-Marquis, Péter Várnai, Erin L Seifert, Gerald W Dorn, Shey-Shing Sheu, György Csordás.

  BACKGROUND: Cardiac contractile function requires high energy from mitochondria, and Ca2+ from the sarcoplasmic reticulum (SR). Via local Ca2+ transfer at close mitochondria-SR contacts, cardiac excitation feedforward regulates mitochondrial ATP production to match surges in demand (excitation-bioenergetics coupling). However, pathological stresses may cause mitochondrial Ca2+ overload, excessive reactive oxygen species production and permeability transition, risking homeostatic collapse and myocyte loss. Excitation-bioenergetics coupling involves mitochondria-SR tethers but the role of tethering in cardiac physiology/pathology is debated. Endogenous tether proteins are multifunctional; therefore, nonselective targets to scrutinize interorganelle linkage. Here, we assessed the physiological/pathological relevance of selective chronic enhancement of cardiac mitochondria-SR tethering.METHODS: We introduced to mice a cardiac muscle-specific engineered tether (linker) transgene with a fluorescent protein core and deployed 2D/3D electron microscopy, biochemical approaches, fluorescence imaging, in vivo and ex vivo cardiac performance monitoring and stress challenges to characterize the linker phenotype.
RESULTS: Expressed in the mature cardiomyocytes, the linker expanded and tightened individual mitochondria-junctional SR contacts; but also evoked a marked remodeling with large dense mitochondrial clusters that excluded dyads. Yet, excitation-bioenergetics coupling remained well-preserved, likely due to more longitudinal mitochondria-dyad contacts and nanotunnelling between mitochondria exposed to junctional SR and those sealed away from junctional SR. Remarkably, the linker decreased female vulnerability to acute massive β-adrenergic stress. It also reduced myocyte death and mitochondrial calcium-overload-associated myocardial impairment in ex vivo ischemia/reperfusion injury.
CONCLUSIONS: We propose that mitochondria-SR/endoplasmic reticulum contacts operate at a structural optimum. Although acute changes in tethering may cause dysfunction, upon chronic enhancement of contacts from early life, adaptive remodeling of the organelles shifts the system to a new, stable structural optimum. This remodeling balances the individually enhanced mitochondrion-junctional SR crosstalk and excitation-bioenergetics coupling, by increasing the connected mitochondrial pool and, presumably, Ca2+/reactive oxygen species capacity, which then improves the resilience to stresses associated with dysregulated hyperactive Ca2+ signaling.

Keywords:  ischemia; myocyte; reperfusion; sarcoplasmic reticulum; transgene

DOI:  https://doi.org/10.1161/CIRCRESAHA.122.321833
bioRxiv. 2023 Mar 29. pii: 2023.03.29.534670. [Epub ahead of print]

Metabolic regulation of misfolded protein import into mitochondria.

Yuhao Wang, Linhao Ruan, Jin Zhu, Xi Zhang, Alexander Chih-Chieh Chang, Alexis Tomaszewski, Rong Li.

Mitochondria are the cellular energy hub and central target of metabolic regulation. Mitochondria also facilitate proteostasis through pathways such as the 'mitochondria as guardian in cytosol' (MAGIC) whereby cytosolic misfolded proteins are imported into and degraded inside mitochondria. In this study, a genome-wide screen in yeast uncovered that Snf1, the yeast AMP-activated protein kinase (AMPK), inhibits the import of misfolded proteins into mitochondria while promoting mitochondrial biogenesis under glucose starvation. We show that this inhibition requires a downstream transcription factor regulating mitochondrial gene expression and is likely to be conferred through substrate competition and mitochondrial import channel selectivity. We further show that Snf1/AMPK activation protects mitochondrial fitness in yeast and human cells under stress induced by misfolded proteins such as those associated with neurodegenerative diseases.

DOI: https://doi.org/10.1101/2023.03.29.534670
ACS Chem Neurosci. 2023 Apr 12.

Cardiolipin and OPA1 Team up for Methamphetamine-Induced Locomotor Activity by Promoting Neuronal Mitochondrial Fusion in the Nucleus Accumbens of Mice.

Liang Wang, Qingfan Wei, Rui Xu, Yaxing Chen, Shu Li, Qian Bu, Ying Zhao, Hongchun Li, Yue Zhao, Linhong Jiang, Yuanyuan Chen, Yanping Dai, Yinglan Zhao, Xiaobo Cen.

  Mitochondria are highly dynamic organelles with coordinated cycles of fission and fusion occurring continuously to satisfy the energy demands in the complex architecture of neurons. How mitochondria contribute to addicted drug-induced adaptable mitochondrial networks and neuroplasticity remains largely unknown. Through liquid chromatography-mass spectrometry-based lipidomics, we first analyzed the alteration of the mitochondrial lipidome of three mouse brain areas in methamphetamine (METH)-induced locomotor activity and conditioned place preference. The results showed that METH remodeled the mitochondrial lipidome of the hippocampus, nucleus accumbens (NAc), and striatum in both models. Notably, mitochondrial hallmark lipid cardiolipin (CL) was specifically increased in the NAc in METH-induced hyperlocomotor activity, which was accompanied by an elongated giant mitochondrial morphology. Moreover, METH significantly boosted mitochondrial respiration and ATP generation as well as the copy number of mitochondrial genome DNA in the NAc. By screening the expressions of mitochondrial dynamin-related proteins, we found that repeated METH significantly upregulated the expression of long-form optic atrophy type 1 (L-OPA1) and enhanced the interaction of L-OPA1 with CL, which may promote mitochondrial fusion in the NAc. On the contrary, neuronal OPA1 depletion in the NAc not only recovered the dysregulated mitochondrial morphology and synaptic vesicle distribution induced by METH but also attenuated the psychomotor effect of METH. Collectively, upregulated CL and OPA1 cooperate to mediate METH-induced adaptation of neuronal mitochondrial dynamics in the NAc, which correlates with the psychomotor effect of METH. These findings propose a potential therapeutic approach for METH addiction by inhibiting neuronal mitochondrial fusion.

Keywords:  OPA1; cardiolipin; locomotor activity; methamphetamine; mitochondrial fusion; nucleus Accumbens

DOI:  https://doi.org/10.1021/acschemneuro.2c00709
Front Microbiol. 2023 ;14 1133773

Mitochondrial nanomotion measured by optical microscopy.

Priyanka Parmar, Maria Ines Villalba, Alexandre Seiji Horii Huber, Aleksandar Kalauzi, Dragana Bartolić, Ksenija Radotić, Ronnie Guy Willaert, Derrick F MacFabe, Sandor Kasas.

  Nanometric scale size oscillations seem to be a fundamental feature of all living organisms on Earth. Their detection usually requires complex and very sensitive devices. However, some recent studies demonstrated that very simple optical microscopes and dedicated image processing software can also fulfill this task. This novel technique, termed as optical nanomotion detection (ONMD), was recently successfully used on yeast cells to conduct rapid antifungal sensitivity tests. In this study, we demonstrate that the ONMD method can monitor motile sub-cellular organelles, such as mitochondria. Here, mitochondrial isolates (from HEK 293 T and Jurkat cells) undergo predictable motility when viewed by ONMD and triggered by mitochondrial toxins, citric acid intermediates, and dietary and bacterial fermentation products (short-chain fatty acids) at various doses and durations. The technique has superior advantages compared to classical methods since it is rapid, possesses a single organelle sensitivity, and is label- and attachment-free.

Keywords:  metabolic substrates; mitochondria; optical nanomotion; rotenone; short chain fatty acids

DOI:  https://doi.org/10.3389/fmicb.2023.1133773
Stem Cell Reports. 2023 Apr 11. pii: S2213-6711(23)00094-2. [Epub ahead of print]18(4): 1030-1047

Scalable generation of sensory neurons from human pluripotent stem cells.

Tao Deng, Vukasin M Jovanovic, Carlos A Tristan, Claire Weber, Pei-Hsuan Chu, Jason Inman, Seungmi Ryu, Yogita Jethmalani, Juliana Ferreira de Sousa, Pinar Ormanoglu, Prisca Twumasi, Chaitali Sen, Jaehoon Shim, Selwyn Jayakar, Han-Xiong Bear Zhang, Sooyeon Jo, Weifeng Yu, Ty C Voss, Anton Simeonov, Bruce P Bean, Clifford J Woolf, Ilyas Singeç.

  Development of new non-addictive analgesics requires advanced strategies to differentiate human pluripotent stem cells (hPSCs) into relevant cell types. Following principles of developmental biology and translational applicability, here we developed an efficient stepwise differentiation method for peptidergic and non-peptidergic nociceptors. By modulating specific cell signaling pathways, hPSCs were first converted into SOX10+ neural crest, followed by differentiation into sensory neurons. Detailed characterization, including ultrastructural analysis, confirmed that the hPSC-derived nociceptors displayed cellular and molecular features comparable to native dorsal root ganglion (DRG) neurons, and expressed high-threshold primary sensory neuron markers, transcription factors, neuropeptides, and over 150 ion channels and receptors relevant for pain research and axonal growth/regeneration studies (e.g., TRPV1, NAV1.7, NAV1.8, TAC1, CALCA, GAP43, DPYSL2, NMNAT2). Moreover, after confirming robust functional activities and differential response to noxious stimuli and specific drugs, a robotic cell culture system was employed to produce large quantities of human sensory neurons, which can be used to develop nociceptor-selective analgesics.

Keywords:  CEPT cocktail; analgesics; cell differentiation; drug testing; iPS cells; neural crest; nociceptor; opioid crisis; pain; sensory neuron

DOI:  https://doi.org/10.1016/j.stemcr.2023.03.006
Science. 2023 Apr 14. 380(6641): 143

Denmark's big tent includes postdocs.

Morten Dall, Maria Hauge Pedersen.

DOI: https://doi.org/10.1126/science.adh4261
Genet Med. 2023 Apr 10. pii: S1098-3600(23)00850-X. [Epub ahead of print] 100837

Clinical implications of conflicting variant interpretations in the cancer genetics clinic.

Elyssa Zukin, Julie O Culver, Yuxi Liu, Yunqi Yang, Charité N Ricker, Rachel Hodan, Duveen Sturgeon, Kerry Kingham, Nicolette M Chun, Courtney Rowe-Teeter, Kathryn Singh, Jason A Zell, Uri Ladabaum, Kevin J McDonnell, James M Ford, Giovanni Parmigiani, Danielle Braun, Allison W Kurian, Stephen B Gruber, Gregory E Idos.

  PURPOSE: To describe the clinical impact of commercial laboratories issuing conflicting classifications of genetic variants.METHODS: Results from 2,000 patients undergoing a multi-gene hereditary cancer panel by a single laboratory were analyzed. Clinically significant discrepancies between the lab provided test reports and other major commercial laboratories were identified, including differences between pathogenic/likely pathogenic (P/LP) and variant of uncertain significance (VUS) classifications, via review of ClinVar archives. For patients carrying a VUS, clinical documentation was assessed for evidence of provider awareness of the conflict.
RESULTS: 50/975 (5.1%) patients with non-negative results carried a variant with a clinically significant conflict, 19 with a P/LP variant reported in APC or MUTYH, and 31 with a VUS reported in CDKN2A, CHEK2, MLH1, MSH2, MUTYH, RAD51C, or TP53. Only 10/28 (36%) patients with a VUS with a clinically significant conflict had a documented discussion by a provider about the conflict. Discrepant counseling strategies were utilized for different patients with the same variant. Among patients with a CDKN2A variant or a monoallelic MUTYH variant, providers were significantly more likely to make recommendations based on the laboratory-reported classification.
CONCLUSION: Our findings highlight the frequency of variant interpretation discrepancies and importance of clinician awareness. Guidance is needed on managing patients with discrepant variants to support accurate risk assessment.

Keywords:  Cancer genetics; ClinVar; VUS; clinical cancer genetics; conflicting variant interpretation; genetic counseling; germline variant; risk assessment; variant interpretation; variant interpretation discrepancy; variant of uncertain significance; variant of unknown significance

DOI:  https://doi.org/10.1016/j.gim.2023.100837
Int J Mol Sci. 2023 Mar 28. pii: 6362. [Epub ahead of print]24(7):

The Role of Alternative Mitophagy in Heart Disease.

Jihoon Nah.

  Autophagy is essential for maintaining cellular homeostasis through bulk degradation of subcellular constituents, including misfolded proteins and dysfunctional organelles. It is generally governed by the proteins Atg5 and Atg7, which are critical regulators of the conventional autophagy pathway. However, recent studies have identified an alternative Atg5/Atg7-independent pathway, i.e., Ulk1- and Rab9-mediated alternative autophagy. More intensive studies have identified its essential role in stress-induced mitochondrial autophagy, also known as mitophagy. Alternative mitophagy plays pathophysiological roles in heart diseases such as myocardial ischemia and pressure overload. Here, this review discusses the established and emerging mechanisms of alternative autophagy/mitophagy that can be applied in therapeutic interventions for heart disorders.

Keywords:  Rab9; alternative autophagy; conventional autophagy; heart disease; mitophagy

DOI:  https://doi.org/10.3390/ijms24076362
Eur J Neurol. 2023 Apr 12.

Clinical and radiological description of 120 pediatric stroke-like episodes.

Chloe Durrleman, David Grevent, Melodie Aubart, Manoelle Kossorotoff, Charles-Joris Roux, Anna Kaminska, Marlene Rio, Giulia Barcia, Nathalie Boddaert, Arnold Munnich, Rima Nabbout, Isabelle Desguerre.

  BACKGROUND: Stroke-like episodes (SLEs) are defined as acute onset of neurological symptoms mimicking a stroke and radiological lesions non-congruent to vascular territory.OBJECTIVE: We aimed to analyze acute clinical and radiological features of SLEs to approach pathophysiology.
METHODS: We performed a monocentre retrospective analysis of 120 SLEs in 60 children over 20 years. Inclusion criteria were: compatible clinical symptoms and stroke-like lesions on brain MRI (performed for all 120 events) with focal hyperintensity on Diffusion-weighted sequence in a non-vascular territory.
RESULTS: Three groups were identified: a first group of mitochondrial diseases (n=22) involving mitochondrial DNA mutations (55%) or nuclear DNA mutations (45%); a second group of other metabolic diseases or epilepsy disorders (n=22), a third group in which no etiology was found despite extensive investigations (n=16). Age at first SLE was younger in metabolic or epilepsy disorders (18 months vs 128 months, p<0.0001) and infectious trigger was more frequent (69% vs 20%, p0.0001). Seizures occurred in 75% of episodes, revealing 50% of SLEs and mainly leading to status epilepticus (90%). Of the 120 MRI scans confirming the diagnosis, 28 were performed within a short and strict 48-hour period and further analyzed to better understand underlying mechanisms. They showed primary cortical hyperintensity (n=28/28) with decreased apparent diffusion coefficient in 52% of the cases. Systematic hyperperfusion was found on Spin Labeling Sequence when available (n=18/18).
CONCLUSION: Clinical and radiological results support the existence of a vicious circle based on two main mechanisms: energy deficit and neuronal hyperexcitability at the origin of SLE.

Keywords:  children; energy deficit; mitochondrial disorder; neuronal hyperexcitability; stroke-like episodes

DOI:  https://doi.org/10.1111/ene.15821
medRxiv. 2023 Mar 29. pii: 2023.03.28.23286936. [Epub ahead of print]

Advancing Understanding of Inequities in Rare Disease Genomics.

Jillian G Serrano, Melanie O'Leary, Grace VanNoy, Ingrid A Holm, Yarden S Fraiman, Heidi L Rehm, Anne O'Donnell-Luria, Monica H Wojcik.

Purpose: Advances in genomic research have led to the diagnosis of rare, early-onset diseases for thousands of individuals. Unfortunately, the benefits of advanced genetic diagnostic technology are not distributed equitably among the population, as has been seen in many other healthcare contexts. Even quantifying and describing inequities in genetic diagnostic yield is challenging due to variation in referrals to clinical genetics practices and other barriers to clinical genetic testing.Methods: The Rare Genomes Project (RGP) at the Broad Institute of MIT and Harvard offers research genome sequencing to individuals with rare disease who remain genetically undiagnosed through direct interaction with the individual or family. This presents an opportunity for diagnosis beyond the clinical context, thus eliminating many barriers to access.
Findings: An initial goal of RGP was to equalize access to genomic sequencing by decoupling testing access from proximity to a major medical center and physician referral. However, our study participants are overwhelmingly non-disadvantaged, as evidenced by their access to specialist care and genetic testing prior to RGP enrollment, and are also predominantly white.
Implications: We therefore describe our novel initiative to diversify RGP enrollment in order to advance equity in rare disease genetic diagnosis and research. In addition to the moral imperative of medical equity, this is also critical in order to fully understand the genomic underpinnings of rare disease. We utilize a mixed methods approach to understand the priorities and values of underrepresented communities, existing disparities, and the obstacles to addressing them: all of which is necessary to promote equity in future genomic medicine initiatives.

DOI: https://doi.org/10.1101/2023.03.28.23286936
Nat Methods. 2023 Apr;20(4): 482

Combined sequencing of genomes and epigenomes.

Lin Tang.

DOI: https://doi.org/10.1038/s41592-023-01856-5
Curr Protoc. 2023 Apr;3(4): e722

Using the Reactome Database.

Karen Rothfels, Marija Milacic, Lisa Matthews, Robin Haw, Cristoffer Sevilla, Marc Gillespie, Ralf Stephan, Chuqiao Gong, Eliot Ragueneau, Bruce May, Veronica Shamovsky, Adam Wright, Joel Weiser, Deidre Beavers, Patrick Conley, Krishna Tiwari, Bijay Jassal, Johannes Griss, Andrea Senff-Ribeiro, Timothy Brunson, Robert Petryszak, Henning Hermjakob, Peter D'Eustachio, Guanming Wu, Lincoln Stein.

  Pathway databases provide descriptions of the roles of proteins, nucleic acids, lipids, carbohydrates, and other molecular entities within their biological cellular contexts. Pathway-centric views of these roles may allow for the discovery of unexpected functional relationships in data such as gene expression profiles and somatic mutation catalogues from tumor cells. For this reason, there is a high demand for high-quality pathway databases and their associated tools. The Reactome project (a collaboration between the Ontario Institute for Cancer Research, New York University Langone Health, the European Bioinformatics Institute, and Oregon Health & Science University) is one such pathway database. Reactome collects detailed information on biological pathways and processes in humans from the primary literature. Reactome content is manually curated, expert-authored, and peer-reviewed and spans the gamut from simple intermediate metabolism to signaling pathways and complex cellular events. This information is supplemented with likely orthologous molecular reactions in mouse, rat, zebrafish, worm, and other model organisms. © 2023 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Browsing a Reactome pathway Basic Protocol 2: Exploring Reactome annotations of disease and drugs Basic Protocol 3: Finding the pathways involving a gene or protein Alternate Protocol 1: Finding the pathways involving a gene or protein using UniProtKB (SwissProt), Ensembl, or Entrez gene identifier Alternate Protocol 2: Using advanced search Basic Protocol 4: Using the Reactome pathway analysis tool to identify statistically overrepresented pathways Basic Protocol 5: Using the Reactome pathway analysis tool to overlay expression data onto Reactome pathway diagrams Basic Protocol 6: Comparing inferred model organism and human pathways using the Species Comparison tool Basic Protocol 7: Comparing tissue-specific expression using the Tissue Distribution tool.

Keywords:  Reactome database; biological pathway; interaction network; pathway analysis; pathway visualization

DOI:  https://doi.org/10.1002/cpz1.722
Orphanet J Rare Dis. 2023 Apr 11. 18(1): 76

Identifying responders to elamipretide in Barth syndrome: Hierarchical clustering for time series data.

Jef Van den Eynde, Bhargava Chinni, Hilary Vernon, W Reid Thompson, Brittany Hornby, Shelby Kutty, Cedric Manlhiot.

  BACKGROUND: Barth syndrome (BTHS) is a rare genetic disease that is characterized by cardiomyopathy, skeletal myopathy, neutropenia, and growth abnormalities and often leads to death in childhood. Recently, elamipretide has been tested as a potential first disease-modifying drug. This study aimed to identify patients with BTHS who may respond to elamipretide, based on continuous physiological measurements acquired through wearable devices.RESULTS: Data from a randomized, double-blind, placebo-controlled crossover trial of 12 patients with BTHS were used, including physiological time series data measured using a wearable device (heart rate, respiratory rate, activity, and posture) and functional scores. The latter included the 6-minute walk test (6MWT), Patient-Reported Outcomes Measurement Information System (PROMIS) fatigue score, SWAY Balance Mobile Application score (SWAY balance score), BTHS Symptom Assessment (BTHS-SA) Total Fatigue score, muscle strength by handheld dynamometry, 5 times sit-and-stand test (5XSST), and monolysocardiolipin to cardiolipin ratio (MLCL:CL). Groups were created through median split of the functional scores into "highest score" and "lowest score", and "best response to elamipretide" and "worst response to elamipretide". Agglomerative hierarchical clustering (AHC) models were implemented to assess whether physiological data could classify patients according to functional status and distinguish non-responders from responders to elamipretide. AHC models clustered patients according to their functional status with accuracies of 60-93%, with the greatest accuracies for 6MWT (93%), PROMIS (87%), and SWAY balance score (80%). Another set of AHC models clustered patients with respect to their response to treatment with elamipretide with perfect accuracy (all 100%).
CONCLUSIONS: In this proof-of-concept study, we demonstrated that continuously acquired physiological measurements from wearable devices can be used to predict functional status and response to treatment among patients with BTHS.

Keywords:  Barth syndrome; Digital health; Hierarchical clustering

DOI:  https://doi.org/10.1186/s13023-023-02676-8