bims-mitdis 2023-03-05 papers

bims-mitdis

Biomed News

on Mitochondrial disorders

Issue of 2023–03–05
forty-one papers selected by
Catalina Vasilescu, Helmholz Munich

The multifaceted mitochondrial OXA insertase.
Mitochondrial pyruvate metabolism regulates the activation of quiescent adult neural stem cells.
Evaluating mitochondrial length, volume, and cristae ultrastructure in rare mouse adult stem cell populations.
Defective COX1 expression in aging mice liver.
Variants in ATP5F1B are associated with dominantly inherited dystonia.
Mitochondrial stress and aging: Lessons from C. elegans.
Mitochondrial proton leak in cardiac aging.
Genotyping Single Nucleotide Polymorphisms in the Mitochondrial Genome by Pyrosequencing.
Role of mitochondrial outer membrane permeabilization during bacterial infection.
Protocol for mitochondrial isolation and sub-cellular localization assay for mitochondrial proteins.
Compartment specific mitochondrial dysfunction in Drosophila knock-in model of ALS reversed by altered gene expression of OXPHOS subunits and pro-fission factor Drp1.
Biotechnological production of reduced and oxidized NAD+ precursors.
Mitochondrial Transport in Axons with Multiple Branching Junctions: A Computational Study.
Kif5a Regulates Mitochondrial Transport in Developing Retinal Ganglion Cells In Vitro.
Analyzing mitochondrial respiration of human induced pluripotent stem cell-derived myeloid progenitors using Seahorse technology.
Case report: malignant hypertension associated with catecholamine excess in a patient with Leigh syndrome.
Standards of NGS Data Sharing and Analysis in Ataxias: Recommendations by the NGS Working Group of the Ataxia Global Initiative.
Ageing is associated with a reduction in markers of mitochondrial energy metabolism in the human epidermis.
The potential role of environmental factors in modulating mitochondrial DNA epigenetic marks.
IGF-1 boosts mitochondrial function by a Ca2+ uptake-dependent mechanism in cultured human and rat cardiomyocytes.
Biallelic pathogenic variants in the mitochondrial form of phosphoenolpyruvate carboxykinase cause peripheral neuropathy.
Impaired mitophagy causes mitochondrial DNA leakage and STING activation in ultraviolet B-irradiated human keratinocytes HaCaT.
An AI-guided screen identifies probucol as an enhancer of mitophagy through modulation of lipid droplets.
High fat diet ameliorates the mitochondrial cardiomyopathy of CHCHD10 mutant mice.
Transgenic NADH dehydrogenase restores oxygen regulation of breathing in mitochondrial complex I-deficient mice.
Optimized protocol for quantification of mitochondrial non-heme and heme iron content in mouse tissues and cultured cells.
Assembly, transfer and fate of mitochondrial iron-sulfur clusters.
Mitochondrial cholesterol: Metabolism and impact on redox biology and disease.
Leukodystrophy Associated with Mitochondrial Complex 1 Deficiency Due to Mutation in NUBPL Gene-An Unusual Follow-Up Finding.
A novel C19orf12 frameshift mutation in a MPAN pedigree impairs mitochondrial function and connectivity leading to neurodegeneration.
Dysfunctional mitochondria accumulate in a skeletal muscle knockout model of Smn1, the causal gene of spinal muscular atrophy.
Mitochondria Dysfunction at the Heart of Viral Myocarditis: Mechanistic Insights and Therapeutic Implications.
Saturated fatty acids increase LPI to reduce FUNDC1 dimerization and stability and mitochondrial function.
Methods and applications for single-cell and spatial multi-omics.
Editorial: Mitochondrial therapy in neurodegeneration.
Computational and mitochondrial functional studies of novel compound heterozygous variants in SPATA5 gene support a causal link with epileptogenic encephalopathy.
Rescuers from the Other Shore: Intercellular Mitochondrial Transfer and Its Implications in Central Nervous System Injury and Diseases.
Inhibition of mitochondrial transcription by the neurotoxin MPP.
Mitochondrial diseases in Hong Kong: prevalence, clinical characteristics and genetic landscape.
Aging and memory are altered by genetically manipulating lactate dehydrogenase in the neurons or glia of flies.
RNA splicing analysis using heterogeneous and large RNA-seq datasets.

Trends Cell Biol. 2023 Feb 28. pii: S0962-8924(23)00020-X. [Epub ahead of print]

The multifaceted mitochondrial OXA insertase.

Bettina Homberg, Peter Rehling, Luis Daniel Cruz-Zaragoza.

  Most mitochondrial proteins are synthesized in the cytosol and transported into mitochondria by protein translocases. Yet, mitochondria contain their own genome and gene expression system, which generates proteins that are inserted in the inner membrane by the oxidase assembly (OXA) insertase. OXA contributes to targeting proteins from both genetic origins. Recent data provides insights into how OXA cooperates with the mitochondrial ribosome during synthesis of mitochondrial-encoded proteins. A picture of OXA emerges in which it coordinates insertion of OXPHOS core subunits and their assembly into protein complexes but also participates in the biogenesis of select imported proteins. These functions position the OXA as a multifunctional protein insertase that facilitates protein transport, assembly, and stability at the inner membrane.

Keywords:  mitochondria; oxidase assembly; oxidative phosphorylation; protein translocation; ribosomes

DOI:  https://doi.org/10.1016/j.tcb.2023.02.001
Sci Adv. 2023 Mar;9(9): eadd5220

Mitochondrial pyruvate metabolism regulates the activation of quiescent adult neural stem cells.

Francesco Petrelli, Valentina Scandella, Sylvie Montessuit, Nicola Zamboni, Jean-Claude Martinou, Marlen Knobloch.

Cellular metabolism is important for adult neural stem/progenitor cell (NSPC) behavior. However, its role in the transition from quiescence to proliferation is not fully understood. We here show that the mitochondrial pyruvate carrier (MPC) plays a crucial and unexpected part in this process. MPC transports pyruvate into mitochondria, linking cytosolic glycolysis to mitochondrial tricarboxylic acid cycle and oxidative phosphorylation. Despite its metabolic key function, the role of MPC in NSPCs has not been addressed. We show that quiescent NSPCs have an active mitochondrial metabolism and express high levels of MPC. Pharmacological MPC inhibition increases aspartate and triggers NSPC activation. Furthermore, genetic Mpc1 ablation in vitro and in vivo also activates NSPCs, which differentiate into mature neurons, leading to overall increased hippocampal neurogenesis in adult and aged mice. These findings highlight the importance of metabolism for NSPC regulation and identify an important pathway through which mitochondrial pyruvate import controls NSPC quiescence and activation.

DOI: https://doi.org/10.1126/sciadv.add5220
STAR Protoc. 2023 Feb 09. pii: S2666-1667(23)00065-5. [Epub ahead of print]4(1): 102107

Evaluating mitochondrial length, volume, and cristae ultrastructure in rare mouse adult stem cell populations.

Matthew Triolo, Steven Wade, Nicole Baker, Mireille Khacho.

  Since changes in mitochondrial morphology regulate key functions of stem cells, it is important to assess their structure under physiological and pathophysiological conditions. Here, we present techniques optimized in rare adult muscle stem cells (MuSCs). For evaluating mitochondrial length and volume within a compact cytoplasmic area in MuSCs on intact myofibers, we describe steps for mitochondrial staining, imaging, and quantification. For evaluating mitochondrial ultrastructure in small cell numbers, we describe steps for agarose embedding and quantification by TEM. For complete details on generation and use of this protocol, please refer to Baker et al. (2022).1.

Keywords:  Cell Biology; Metabolism; Microscopy; Stem Cells

DOI:  https://doi.org/10.1016/j.xpro.2023.102107
Biol Open. 2023 Mar 15. pii: bio059844. [Epub ahead of print]12(3):

Defective COX1 expression in aging mice liver.

Steffen Witte, Angela Boshnakovska, Metin Özdemir, Arpita Chowdhury, Peter Rehling, Abhishek Aich.

  Mitochondrial defects are associated with aging processes and age-related diseases, including cardiovascular diseases, neurodegenerative diseases and cancer. In addition, some recent studies suggest mild mitochondrial dysfunctions appear to be associated with longer lifespans. In this context, liver tissue is considered to be largely resilient to aging and mitochondrial dysfunction. Yet, in recent years studies report dysregulation of mitochondrial function and nutrient sensing pathways in ageing livers. Therefore, we analyzed the effects of the aging process on mitochondrial gene expression in liver using wildtype C57BL/6N mice. In our analyses, we observed alteration in mitochondrial energy metabolism with age. To assess if defects in mitochondrial gene expression are linked to this decline, we applied a Nanopore sequencing based approach for mitochondrial transcriptomics. Our analyses show that a decrease of the Cox1 transcript correlates with reduced respiratory complex IV activity in older mice livers.

Keywords:  Ageing; Mitochondria; Nanopore; Transcriptomics

DOI:  https://doi.org/10.1242/bio.059844
Brain. 2023 Mar 01. pii: awad068. [Epub ahead of print]

Variants in ATP5F1B are associated with dominantly inherited dystonia.

Alessia Nasca, Niccolò E Mencacci, Federica Invernizzi, Michael Zech, Ignacio J Keller Sarmiento, Andrea Legati, Chiara Frascarelli, Bernabe I Bustos, Luigi M Romito, Dimitri Krainc, Juliane Winkelmann, Miryam Carecchio, Nardo Nardocci, Giovanna Zorzi, Holger Prokisch, Steven J Lubbe, Barbara Garavaglia, Daniele Ghezzi.

  ATP5F1B is a subunit of the mitochondrial ATP synthase or complex V of the mitochondrial respiratory chain. Pathogenic variants in nuclear genes encoding assembly factors or structural subunits are associated with complex V deficiency, typically characterized by autosomal recessive inheritance and multisystem phenotypes. Movement disorders have been described in a subset of cases carrying autosomal dominant variants in structural subunits genes ATP5F1A and ATP5MC3. Here, we report the identification of two different ATP5F1B missense variants (c.1000A>C; p.Thr334Pro and c.1445T>C; p.Val482Ala) segregating with early-onset isolated dystonia in two families, both with autosomal dominant mode of inheritance and incomplete penetrance. Functional studies in mutant fibroblasts revealed no decrease of ATP5F1B protein amount but severe reduction of complex V activity and impaired mitochondrial membrane potential, suggesting a dominant-negative effect. In conclusion, our study describes a new candidate gene associated with isolated dystonia and confirms that heterozygous variants in genes encoding subunits of the mitochondrial ATP synthase may cause autosomal dominant isolated dystonia with incomplete penetrance, likely through a dominant-negative mechanism.

Keywords:  ATP5F1B; case report; dystonia; incomplete penetrance; mitochondrial ATP synthase

DOI:  https://doi.org/10.1093/brain/awad068
Semin Cell Dev Biol. 2023 Feb 28. pii: S1084-9521(23)00038-1. [Epub ahead of print]

Mitochondrial stress and aging: Lessons from C. elegans.

Peng X Chen, Leyuan Zhang, Di Chen, Ye Tian.

  Aging is accompanied by a progressive decline in mitochondrial function, which in turn contributes to a variety of age-related diseases. Counterintuitively, a growing number of studies have found that disruption of mitochondrial function often leads to increased lifespan. This seemingly contradictory observation has inspired extensive research into genetic pathways underlying the mitochondrial basis of aging, particularly within the model organism Caenorhabditis elegans. The complex and antagonistic roles of mitochondria in the aging process have altered the view of mitochondria, which not only serve as simple bioenergetic factories but also as signaling platforms for the maintenance of cellular homeostasis and organismal health. Here, we review the contributions of C. elegans to our understanding of mitochondrial function in the aging process over the past decades. In addition, we explore how these insights may promote future research of mitochondrial-targeted strategies in higher organisms to potentially slow aging and delay age-related disease progression.

Keywords:  Aging; C. elegans; Longevity; Mitochondrial stress; Mitokine

DOI:  https://doi.org/10.1016/j.semcdb.2023.02.010
Geroscience. 2023 Mar 01.

Mitochondrial proton leak in cardiac aging.

Xingyun Qi, Nancy J Rusch, Jiaojiao Fan, Christoph J Mora, Lixin Xie, Shengyu Mu, Peter S Rabinovitch, Huiliang Zhang.

  Age-associated diseases are becoming progressively more prevalent, reflecting the increased lifespan of the world's population. However, the fundamental mechanisms of physiologic aging are poorly understood, and in particular, the molecular pathways that mediate cardiac aging and its associated dysfunction are unclear. Here, we focus on certain ion flux abnormalities of the mitochondria that may contribute to cardiac aging and age-related heart failure. Using oxidative phosphorylation, mitochondria pump protons from the matrix to the intermembrane space to generate a proton gradient across the inner membrane. The protons are returned to the matrix by the ATPase complex within the membrane to generate ATP. However, a portion of protons leak back to the matrix and do not drive ATP production, and this event is called proton leak or uncoupling. Accumulating evidence suggests that mitochondrial proton leak is increased in the cardiac myocytes of aged hearts. In this mini-review, we discuss the measurement methods and major sites of mitochondrial proton leak with an emphasis on the adenine nucleotide transporter 1 (ANT1), and explore the possibility of inhibiting augmented mitochondrial proton leak as a therapeutic intervention to mitigate cardiac aging.

Keywords:  Adenine nucleotide transporter 1; Cardiac aging; Heart failure; Mitochondria; Pleiotropic role; Proton leak

DOI:  https://doi.org/10.1007/s11357-023-00757-x
J Vis Exp. 2023 Feb 10.

Genotyping Single Nucleotide Polymorphisms in the Mitochondrial Genome by Pyrosequencing.

Pavel A Nash, Pedro Silva-Pinheiro, Michal A Minczuk.

Mutations in the mitochondrial genome (mtDNA) have been associated with maternally inherited genetic diseases. However, interest in mtDNA polymorphisms has increased in recent years due to the recently developed ability to produce models by mtDNA mutagenesis and a new appreciation of the association between mitochondrial genetic aberrations and common age-related diseases such as cancer, diabetes, and dementia. Pyrosequencing is a sequencing-by-synthesis technique that is widely employed across the mitochondrial field for routine genotyping experiments. Its relative affordability when compared to massive parallel sequencing methods and ease of implementation make it an invaluable technique in the field of mitochondrial genetics, allowing for the rapid quantification of heteroplasmy with increased flexibility. Despite the practicality of this method, its implementation as a means of mtDNA genotyping requires the observation of certain guidelines, specifically to avoid certain biases of biological or technical origin. This protocol outlines the necessary steps and precautions in designing and implementing pyrosequencing assays for use in the context of heteroplasmy measurement.

DOI: https://doi.org/10.3791/64361
Int Rev Cell Mol Biol. 2023 ;pii: S1937-6448(22)00136-8. [Epub ahead of print]374 83-127

Role of mitochondrial outer membrane permeabilization during bacterial infection.

Collins Waguia Kontchou, Georg Häcker.

  Beyond the initial 'powerhouse' view, mitochondria have numerous functions in their mammalian cell and contribute to many physiological processes, and many of these we understand only partially. The control of apoptosis by mitochondria is firmly established. Many questions remain however how this function is embedded into physiology, and how other signaling pathways regulate mitochondrial apoptosis; the interplay of bacteria with the mitochondrial apoptosis pathway is one such example. The outer mitochondrial membrane regulates both import into mitochondria and the release of intermembrane, and in some situations also matrix components from mitochondria, and these mitochondrial components can have signaling function in the cytosol. One function is the induction of apoptotic cell death. An exciting, more recently discovered function is the regulation of inflammation. Mitochondrial molecules, both proteins and nucleic acids, have inflammatory activity when released from mitochondria, an activity whose regulation is intertwined with the activation of apoptotic caspases. Bacterial infection can have more general effects on mitochondrial apoptosis-regulation, through effects on host transcription and other pathways, such as signals controlled by pattern recognition. Some specialized bacteria have products that more specifically regulate signaling to the outer mitochondrial membrane, and to apoptosis; both pro- and anti-apoptotic mechanisms have been reported. Among the intriguing recent findings in this area are signaling contributions of porins and the sub-lethal release of intermembrane constituents. We will here review the literature and place the new developments into the established context of mitochondrial signaling during the contact of bacterial pathogens with human cells.

Keywords:  Anti-apoptotic; Bacterial infection; Cytochrome c; Host cell; Immune response; Macrophage; Mitochondria; Outer mitochondrial membrane; Pro-apoptotic; Receptors; Second mitochondria-derived activator of caspase (Smac)

DOI:  https://doi.org/10.1016/bs.ircmb.2022.10.002
STAR Protoc. 2023 Feb 03. pii: S2666-1667(23)00046-1. [Epub ahead of print]4(1): 102088

Protocol for mitochondrial isolation and sub-cellular localization assay for mitochondrial proteins.

Danyi Zhou, Sheng Zhong, Xinyu Han, Dandan Liu, Hezhi Fang, Ya Wang.

  Here, we provide a protocol to isolate mitochondria from cultured cells and extract differently located mitochondrial proteins. We detail steps to separate both integral and peripheral membrane proteins from soluble proteins using sonication. We describe the separation of integral membrane proteins from the peripheral membrane and soluble proteins using sodium carbonate extraction. Furthermore, we detail the use of proteinase K and Triton X-100 to distinguish outer membrane proteins from mitochondrial proteins.

Keywords:  Cell Membrane; Cell culture; Cell separation/fractionation; Protein Biochemistry

DOI:  https://doi.org/10.1016/j.xpro.2023.102088
Mol Cell Neurosci. 2023 Mar 01. pii: S1044-7431(23)00028-3. [Epub ahead of print] 103834

Compartment specific mitochondrial dysfunction in Drosophila knock-in model of ALS reversed by altered gene expression of OXPHOS subunits and pro-fission factor Drp1.

Y Nemtsova, B L Steinert, K A Wharton.

  Amyotrophic Lateral Sclerosis (ALS) is a fatal multisystem neurodegenerative disease, characterized by a loss in motor function. ALS is genetically diverse, with mutations in genes ranging from those regulating RNA metabolism, like TAR DNA-binding protein (TDP-43) and Fused in sarcoma (FUS), to those that act to maintain cellular redox homeostasis, like superoxide dismutase 1 (SOD1). Although varied in genetic origin, pathogenic and clinical commonalities are clearly evident between cases of ALS. Defects in mitochondria is one such common pathology, thought to occur prior to, rather than as a consequence of symptom onset, making these organelles a promising therapeutic target for ALS, as well as other neurodegenerative diseases. Depending on the homeostatic needs of neurons throughout life, mitochondria are normally shuttled to different subcellular compartments to regulate metabolite and energy production, lipid metabolism, and buffer calcium. While originally considered a motor neuron disease due to the dramatic loss in motor function accompanied by motor neuron cell death in ALS patients, many studies have now implicated non-motor neurons and glial cells alike. Defects in non-motor neuron cell types often preceed motor neuron death suggesting their dysfunction may initiate and/or facilitate the decline in motor neuron health. Here, we investigate mitochondria in a Drosophila Sod1 knock-in model of ALS. In depth, in vivo, examination reveals mitochondrial dysfunction evident prior to onset of motor neuron degeneration. Genetically encoded redox biosensors identify a general disruption in the electron transport chain (ETC). Compartment specific abnormalities in mitochondrial morphology is observed in diseased sensory neurons, accompanied by no apparent defects in the axonal transport machinery, but instead an increase in mitophagy in synaptic regions. The decrease in networked mitochondria at the synapse is reversed upon downregulation of the pro-fission factor Drp1. Furthermore, altered expression of specific OXPHOS subunits reverses ALS-associated defects in mitochondrial morphology and function.

Keywords:  ALS; Drosophila; Mitochondrial dysfunction; Neurodegeneration; Sensory neurons

DOI:  https://doi.org/10.1016/j.mcn.2023.103834
Food Res Int. 2023 Mar;pii: S0963-9969(23)00105-9. [Epub ahead of print]165 112560

Biotechnological production of reduced and oxidized NAD+ precursors.

Rubén Zapata-Pérez, Antonio Ginés García-Saura, Angelique M L Scantlebery, Bauke V Schomakers, Rubén Rabadán-Ros, Michel van Weeghel, Riekelt H Houtkooper, Álvaro Sánchez-Ferrer.

  Dysregulation of nicotinamide adenine dinucleotide (NAD+) homeostasis by increased activity of NAD+ consumers or reduced NAD+ biosynthesis plays an important role in the onset of prevalent, often age-related, diseases, such as diabetes, neuropathies or nephropathies. To counteract such dysregulation, NAD+ replenishment strategies can be used. Among these, administration of vitamin B3 derivatives (NAD+ precursors) has garnered attention in recent years. However, the high market price of these compounds and their limited availability, pose important limitations to their use in nutritional or biomedical applications. To overcome these limitations, we have designed an enzymatic method for the synthesis and purification of (1) the oxidized NAD+ precursors nicotinamide mononucleotide (NMN) and nicotinamide riboside (NR), (2) their reduced forms NMNH and NRH, and (3) their deaminated forms nicotinic acid mononucleotide (NaMN) and nicotinic acid riboside (NaR). Starting from NAD+ or NADH as substrates, we use a combination of three highly overexpressed soluble recombinant enzymes; (a) a NAD+ pyrophosphatase, (b) an NMN deamidase, and (c) a 5'-nucleotidase, to produce these six precursors. Finally, we validate the activity of the enzymatically produced molecules as NAD+ enhancers in cell culture.

Keywords:  Enzymatic synthesis; NAD precursor; NaMN

DOI:  https://doi.org/10.1016/j.foodres.2023.112560
bioRxiv. 2023 Feb 23. pii: 2023.02.22.529604. [Epub ahead of print]

Mitochondrial Transport in Axons with Multiple Branching Junctions: A Computational Study.

Ivan A Kuznetsov, Andrey V Kuznetsov.

We explore the impact of multiple branching junctions in axons on the mean age of mitochondria and their age density distributions in demand sites. The study looked at mitochondrial concentration, mean age, and age density distribution in relation to the distance from the soma. We developed models for a symmetric axon containing 14 demand sites and an asymmetric axon containing 10 demand sites. We examined how the concentration of mitochondria changes when an axon is divided into two branches at the branching junction. We also studied whether mitochondria concentrations in the branches are affected by what portion of mitochondrial flux goes into the upper branch and what portion goes into the lower branch. Additionally, we explored whether the distributions of mitochondria mean age and age density in branching axons are affected by how the mitochondria flux splits at the branching junction. Our findings elucidate the effects of axonal branching on mitochondria age. Mitochondria aging is the focus of this study as recent research suggests it may be involved in neurodegenerative disorders.

DOI: https://doi.org/10.1101/2023.02.22.529604
Invest Ophthalmol Vis Sci. 2023 Mar 01. 64(3): 4

Kif5a Regulates Mitochondrial Transport in Developing Retinal Ganglion Cells In Vitro.

Satoshi Yokota, Sahil H Shah, Emma Lee Huie, Runxia Rain Wen, Ziming Luo, Jeffrey L Goldberg.

Purpose: Axon transport of organelles and neurotrophic factors is necessary for maintaining cellular function and survival of retinal ganglion cells (RGCs). However, it is not clear how trafficking of mitochondria, essential for RGC growth and maturation, changes during RGC development. The purpose of this study was to understand the dynamics and regulation of mitochondrial transport during RGC maturation using acutely purified RGCs as a model system.
Methods: Primary RGCs were immunopanned from rats of either sex during three stages of development. MitoTracker dye and live-cell imaging were used to quantify mitochondrial motility. Analysis of single-cell RNA sequencing was used to identify Kinesin family member 5A (Kif5a) as a relevant motor candidate for mitochondrial transport. Kif5a expression was manipulated with either short hairpin RNA (shRNA) or exogenous expression adeno-associated virus viral vectors.
Results: Anterograde and retrograde mitochondrial trafficking and motility decreased through RGC development. Similarly, the expression of Kif5a, a motor protein that transports mitochondria, also decreased during development. Kif5a knockdown decreased anterograde mitochondrial transport, while Kif5a expression increased general mitochondrial motility and anterograde mitochondrial transport.
Conclusions: Our results suggested that Kif5a directly regulates mitochondrial axonal transport in developing RGCs. Future work exploring the role of Kif5a in vivo in RGCs is indicated.

DOI: https://doi.org/10.1167/iovs.64.3.4
STAR Protoc. 2023 Jan 28. pii: S2666-1667(23)00031-X. [Epub ahead of print]4(1): 102073

Analyzing mitochondrial respiration of human induced pluripotent stem cell-derived myeloid progenitors using Seahorse technology.

Yanxin Fan, Yoko Mizoguchi, Megumi Tatematsu, Monika I Linder, Stephanie Frenz, Jongsu Choi, Christoph Klein.

  Mitochondrial metabolism is critical in hematopoietic stem cell maintenance and differentiation. Here, we present a step-by-step protocol to efficiently differentiate human induced pluripotent stem cells into myeloid progenitors by a robust feeder- and serum-free system. Furthermore, we provide a protocol to subsequently assess mitochondrial function in iPSC-derived myeloid progenitors. We comprehensively describe a protocol to analyze and to quantify key parameters of mitochondrial respiration of iPSC-derived myeloid progenitors by the Seahorse XFe96 Analyzer. Additionally, our protocol includes extensive troubleshooting suggestions. For complete details on the use and execution of this protocol, please refer to Fan et al. (2022).1.

Keywords:  Cell Biology; Cell Differentiation; Immunology; Metabolism; Stem Cells

DOI:  https://doi.org/10.1016/j.xpro.2023.102073
Clin Hypertens. 2023 Mar 01. 29(1): 7

Case report: malignant hypertension associated with catecholamine excess in a patient with Leigh syndrome.

Ana Solis, Joshua Shimony, Marwan Shinawi, Kevin T Barton.

   BACKGROUND: Leigh syndrome is a progressive neurodegenerative mitochondrial disorder caused by multiple genetic etiologies with multisystemic involvement that mostly affecting the central nervous system with high rate of premature mortality.
CASE PRESENTATION: We present a 3-year, 10 month-old female patient with Leigh syndrome complicated by renal tubular acidosis, hypertension, gross motor delay, who presented with hypertensive emergency, persistent tachycardia, insomnia and irritability. Her previous genetic workup revealed a pathogenic variant in the MT-ND5 gene designated as m.13513G > A;p.Asp393Asn with a heteroplasmy of 69%. She presented acutely with malignant hypertension requiring intensive care unit admission. Her acute evaluation revealed elevated serum and urine catecholamines, without an identifiable catecholamine-secreting tumor. After extensive evaluation for secondary causes, she was ultimately found to have progression of her disease with new infarctions in her medulla, pons, and basal ganglia as the most likely etiology of her hypertension. She was discharged home with clonidine, amlodipine and atenolol for hypertension management. This report highlights the need to recognize possible autonomic dysfunction in mitochondrial disease and illustrates the challenges for accurate and prompt diagnosis and subsequent management of the associated manifestations. This association between catecholamine induced autonomic dysfunction and Leigh syndrome has been previously reported only once with MT-ND5 mutation.
CONCLUSIONS: Elevated catecholamines with malignant secondary hypertension may be unique to this specific mutation or may be a previously unrecognized feature of Leigh syndrome and other mitochondrial complex I deficient syndromes. As such, patients with Leigh syndrome who present with malignant hypertension should be treated without the need for extensive work-up for catecholamine-secreting tumors.

Keywords:  Hypertension; Leigh syndrome; Nephrology; Pediatrics; Secondary hypertension

DOI:  https://doi.org/10.1186/s40885-022-00231-4
Cerebellum. 2023 Mar 04.

Standards of NGS Data Sharing and Analysis in Ataxias: Recommendations by the NGS Working Group of the Ataxia Global Initiative.

AGI Ataxia NGS genomics, platforms Working Group

  The Ataxia Global Initiative (AGI) is a worldwide multi-stakeholder research platform to systematically enhance trial-readiness in degenerative ataxias. The next-generation sequencing (NGS) working group of the AGI aims to improve methods, platforms, and international standards for ataxia NGS analysis and data sharing, ultimately allowing to increase the number of genetically ataxia patients amenable for natural history and treatment trials. Despite extensive implementation of NGS for ataxia patients in clinical and research settings, the diagnostic gap remains sizeable, as approximately 50% of patients with hereditary ataxia remain genetically undiagnosed. One current shortcoming is the fragmentation of patients and NGS datasets on different analysis platforms and databases around the world. The AGI NGS working group in collaboration with the AGI associated research platforms-CAGC, GENESIS, and RD-Connect GPAP-provides clinicians and scientists access to user-friendly and adaptable interfaces to analyze genome-scale patient data. These platforms also foster collaboration within the ataxia community. These efforts and tools have led to the diagnosis of > 500 ataxia patients and the discovery of > 30 novel ataxia genes. Here, the AGI NGS working group presents their consensus recommendations for NGS data sharing initiatives in the ataxia field, focusing on harmonized NGS variant analysis and standardized clinical and metadata collection, combined with collaborative data and analysis tool sharing across platforms.

Keywords:  Cerebellar ataxia; Consensus; Genomics; High-throughput nucleotide sequencing; Information dissemination

DOI:  https://doi.org/10.1007/s12311-023-01537-1
Exp Dermatol. 2023 Feb 27.

Ageing is associated with a reduction in markers of mitochondrial energy metabolism in the human epidermis.

Silvia Vidali, René G Feichtinger, Michael Emberger, Susanne Maria Brunner, Stefanie Gaisbauer, Thomas Blatt, William J Smiles, Christina Kreutzer, Julia M Weise, Barbara Kofler.

  The decline of mitochondrial function throughout the lifespan is directly linked to the development of ageing phenotypes of the skin. Here, we assessed alterations in markers of epidermal mitochondrial energy metabolism as a function of skin age. Human skin samples from distinct anatomical regions were obtained during routine dermatological surgery from 21 young (27.6 ± 1.71 year) and 22 old (76.2 ± 1.73 year) donors. Sections of skin samples were analysed by immunohistochemistry for mitochondrial subunits of each electron transport chain complex (I-V)/oxidative phosphorylation (OXPHOS), as well as proteins serving as a marker of mitochondrial mass (VDAC1) and the regulation of DNA transcription (TFAM). Staining intensities of ATP5F1A (comprising complex V) and TFAM in the epidermis of older subjects were significantly decreased compared with younger donors. Moreover, these effects were independent of UV exposure of the stained skin section. Overall, we demonstrate that ageing is associated with reduced protein levels of complex V of the mitochondrial respiratory chain and TFAM. These alterations may impair essential mitochondrial functions, exacerbating the cutaneous ageing process.

Keywords:  OXPHOS; ageing; epidermis; mitochondria; skin

DOI:  https://doi.org/10.1111/exd.14778
Vitam Horm. 2023 ;pii: S0083-6729(23)00011-0. [Epub ahead of print]122 107-145

The potential role of environmental factors in modulating mitochondrial DNA epigenetic marks.

Brad A Ryva, Zheng Zhou, Jaclyn M Goodrich, Rita S Strakovsky.

  Many studies implicate mitochondrial dysfunction in the development and progression of numerous chronic diseases. Mitochondria are responsible for most cellular energy production, and unlike other cytoplasmic organelles, mitochondria contain their own genome. Most research to date, through investigating mitochondrial DNA copy number, has focused on larger structural changes or alterations to the entire mitochondrial genome and their role in human disease. Using these methods, mitochondrial dysfunction has been linked to cancers, cardiovascular disease, and metabolic health. However, like the nuclear genome, the mitochondrial genome may experience epigenetic alterations, including DNA methylation that may partially explain some of the health effects of various exposures. Recently, there has been a movement to understand human health and disease within the context of the exposome, which aims to describe and quantify the entirety of all exposures people encounter throughout their lives. These include, among others, environmental pollutants, occupational exposures, heavy metals, and lifestyle and behavioral factors. In this chapter, we summarize the current research on mitochondria and human health, provide an overview of the current knowledge on mitochondrial epigenetics, and describe the experimental and epidemiologic studies that have investigated particular exposures and their relationships with mitochondrial epigenetic modifications. We conclude the chapter with suggestions for future directions in epidemiologic and experimental research that is needed to advance the growing field of mitochondrial epigenetics.

Keywords:  Endocrine disruptor; Environmental exposure; Epigenetics; Exposome; Methylation; Mitochondria; Particulate matter

DOI:  https://doi.org/10.1016/bs.vh.2023.01.011
Front Physiol. 2023 ;14 1106662

IGF-1 boosts mitochondrial function by a Ca2+ uptake-dependent mechanism in cultured human and rat cardiomyocytes.

Pablo Sánchez-Aguilera, Camila López-Crisosto, Ignacio Norambuena-Soto, Christian Penannen, Jumo Zhu, Nils Bomer, Matijn F Hoes, Peter Van Der Meer, Mario Chiong, B Daan Westenbrink, Sergio Lavandero.

  A physiological increase in cardiac workload results in adaptive cardiac remodeling, characterized by increased oxidative metabolism and improvements in cardiac performance. Insulin-like growth factor-1 (IGF-1) has been identified as a critical regulator of physiological cardiac growth, but its precise role in cardiometabolic adaptations to physiological stress remains unresolved. Mitochondrial calcium (Ca2+) handling has been proposed to be required for sustaining key mitochondrial dehydrogenase activity and energy production during increased workload conditions, thus ensuring the adaptive cardiac response. We hypothesized that IGF-1 enhances mitochondrial energy production through a Ca2+-dependent mechanism to ensure adaptive cardiomyocyte growth. We found that stimulation with IGF-1 resulted in increased mitochondrial Ca2+ uptake in neonatal rat ventricular myocytes and human embryonic stem cell-derived cardiomyocytes, estimated by fluorescence microscopy and indirectly by a reduction in the pyruvate dehydrogenase phosphorylation. We showed that IGF-1 modulated the expression of mitochondrial Ca2+ uniporter (MCU) complex subunits and increased the mitochondrial membrane potential; consistent with higher MCU-mediated Ca2+ transport. Finally, we showed that IGF-1 improved mitochondrial respiration through a mechanism dependent on MCU-mediated Ca2+ transport. In conclusion, IGF-1-induced mitochondrial Ca2+ uptake is required to boost oxidative metabolism during cardiomyocyte adaptive growth.

Keywords:  MCU complex; human embryonic stem cell derived-cardiomyocytes (hES-CMs); insulin-like growth factor 1 (IGF-1); mitochondrial calcium handling; neonatal rat ventricular myocytes (NRVMs); physiological cardiac hypertrophy

DOI:  https://doi.org/10.3389/fphys.2023.1106662
HGG Adv. 2023 Apr 13. 4(2): 100182

Biallelic pathogenic variants in the mitochondrial form of phosphoenolpyruvate carboxykinase cause peripheral neuropathy.

Neal Sondheimer, Alberto Aleman, Jessie Cameron, Hernan Gonorazky, Nesrin Sabha, Paula Oliveira, Kimberly Amburgey, Azizia Wahedi, Dahai Wang, Michael Shy, James J Dowling.

  Phosphoenolpyruvate carboxykinase (PCK) plays a critical role in cytosolic gluconeogenesis, and defects in PCK1 cause a fasting-aggravated metabolic disease with hypoglycemia and lactic acidosis. However, there are two genes encoding PCK, and the role of the mitochondrial resident PCK (encoded by PCK2) is unclear, since gluconeogenesis is cytosolic. We identified three patients in two families with biallelic variants in PCK2. One has compound heterozygous variants (p.Ser23Ter/p.Pro170Leu), and the other two (siblings) have homozygous p.Arg193Ter variation. All three patients have weakness and abnormal gait, an absence of PCK2 protein, and profound reduction in PCK2 activity in fibroblasts, but no obvious metabolic phenotype. Nerve conduction studies showed reduced conduction velocities with temporal dispersion and conduction block compatible with a demyelinating peripheral neuropathy. To validate the association between PCK2 variants and clinical disease, we generated a mouse knockout model of PCK2 deficiency. The animals present abnormal nerve conduction studies and peripheral nerve pathology, corroborating the human phenotype. In total, we conclude that biallelic variants in PCK2 cause a neurogenetic disorder featuring abnormal gait and peripheral neuropathy.

Keywords:  PCK2; mitochondria; neurogenetic; peripheral neuropathy

DOI:  https://doi.org/10.1016/j.xhgg.2023.100182
Arch Biochem Biophys. 2023 Feb 25. pii: S0003-9861(23)00052-8. [Epub ahead of print]737 109553

Impaired mitophagy causes mitochondrial DNA leakage and STING activation in ultraviolet B-irradiated human keratinocytes HaCaT.

Can Li, Yuying Zhu, Weiwei Liu, Wendie Xiang, Sijun He, Toshihiko Hayashi, Kazunori Mizuno, Shunji Hattori, Hitomi Fujisaki, Takashi Ikejima.

  Ultraviolet B (UVB) irradiation causes skin damages. In this study, we focus on the involvement of mitochondrial disorders in UVB injury. Surprisingly, UVB irradiation increases the amounts of mitochondria in human immortalized keratinocytes HaCaT. However, further analysis shows that ATP levels decreased by UVB treatment in accordance with the collapse of mitochondrial membrane potential (MMP), suggesting an accumulation of dysfunctional mitochondria in UVB-irradiated HaCaT cells. Mitophagy, mainly mediated by PINK1 and parkin, is critical for the elimination of damaged mitochondria. Western blot results show that the levels of both PINK1 and parkin are decreased in UVB-irradiated cells, indicating the impairment of mitophagy. Silencing the expression of PINK1 or parkin by transfection of siRNA shows essentially the same damage to the cells as UVB irradiation does, including increased mitochondrial amount, decreased MMP and ATP production, and enhanced apoptosis, evidencing that repression of PINK1/parkin-mediated mitophagy plays a primary cause of UVB-caused cells damages. We previously found that HaCaT cells exposed to UVB showed activation of the cGAS-STING pathway and apoptosis. Here, silencing PINK1 or parkin also increases the protein levels of cGAS and STING, facilitates nuclear accumulation of NF-κB, and promotes the transcription of IFNβ, suggesting for the activation of STING pathway. Mitophagy impairment either by UVB-irradiation or by PINK1/parkin silencing initiates caspase-3-mediated apoptosis, as shown by the activation of caspase-3 and cleavage of PARP, as well as the increase of Hoechst-positive stained cells and Annexin V-positive cells. Further studies find that Bax-mediated permeabilization of mitochondrial membrane is critical for cell apoptosis, as well as the cytosolic leakage of mtDNA in UVB-treated cells, which results in cGAS-STING activation, and these processes are negatively-regulated by PINK1/parkin-mediated mitophagy. This study reveals the involvement of dysfunctional mitochondria due to impaired mitophagy in the damaging effect of UVB irradiation on HaCaT cells. Restoring the mitophagy has the potential to be developed as a new strategy to protect skin from UVB damages.

Keywords:  Apoptosis; Mitochondrial DNA; PINK1; Parkin; STING; UVB

DOI:  https://doi.org/10.1016/j.abb.2023.109553
PLoS Biol. 2023 Mar;21(3): e3001977

An AI-guided screen identifies probucol as an enhancer of mitophagy through modulation of lipid droplets.

Natalia Moskal, Naomi P Visanji, Olena Gorbenko, Vijay Narasimhan, Hannah Tyrrell, Jess Nash, Peter N Lewis, G Angus McQuibban.

Failures in mitophagy, a process by which damaged mitochondria are cleared, results in neurodegeneration, while enhancing mitophagy promotes the survival of dopaminergic neurons. Using an artificial intelligence platform, we employed a natural language processing approach to evaluate the semantic similarity of candidate molecules to a set of well-established mitophagy enhancers. Top candidates were screened in a cell-based mitochondrial clearance assay. Probucol, a lipid-lowering drug, was validated across several orthogonal mitophagy assays. In vivo, probucol improved survival, locomotor function, and dopaminergic neuron loss in zebrafish and fly models of mitochondrial damage. Probucol functioned independently of PINK1/Parkin, but its effects on mitophagy and in vivo depended on ABCA1, which negatively regulated mitophagy following mitochondrial damage. Autophagosome and lysosomal markers were elevated by probucol treatment in addition to increased contact between lipid droplets (LDs) and mitochondria. Conversely, LD expansion, which occurs following mitochondrial damage, was suppressed by probucol and probucol-mediated mitophagy enhancement required LDs. Probucol-mediated LD dynamics changes may prime the cell for a more efficient mitophagic response to mitochondrial damage.

DOI: https://doi.org/10.1371/journal.pbio.3001977
bioRxiv. 2023 Feb 22. pii: 2023.02.22.529577. [Epub ahead of print]

High fat diet ameliorates the mitochondrial cardiomyopathy of CHCHD10 mutant mice.

Nneka Southwell, Dazhi Zhao, Nicole M Sayles, Jalia Dash, Keigo Fujita, Marilena Dâ Aurelio, Giovanni Manfredi, Hibiki Kawamata.

Mutations in CHCHD10, a mitochondrial protein with still undefined function, are associated with dominant multi-system mitochondrial diseases. CHCHD10 knock-in mice harboring a heterozygous S55L mutation (equivalent to the human pathogenic S59L mutation) develop a fatal mitochondrial cardiomyopathy. The heart of S55L knock-in mice shows extensive metabolic rewiring triggered by proteotoxic mitochondrial integrated stress response (mtISR). In the mutant heart, mtISR initiates well before the onset of mild bioenergetic impairments and is associated with a shift from fatty acid oxidation to glycolytic metabolism and widespread metabolic imbalance. We tested therapeutic interventions to counteract the metabolic rewiring and ameliorate the metabolic imbalance. Heterozygous S55L mice were subjected to chronic high fat diet (HFD) to decrease insulin sensitivity and glucose uptake and enhance fatty acid utilization in the heart. Metabolomics and gene expression profiles demonstrated that HFD achieved an increase of fatty acid utilization in the heart accompanied by a decrease in cardiomyopathy markers. Surprisingly, HFD also decreased the accumulation of aggregated CHCHD10 in the S55L heart. Importantly, HFD increased the survival of mutant female mice exposed to acceleration of the mitochondrial cardiomyopathy associated with pregnancy. Our findings indicate that metabolic alterations can be effectively targeted for therapeutic intervention in mitochondrial cardiomyopathies associated with proteotoxic stress.

DOI: https://doi.org/10.1101/2023.02.22.529577
Nat Commun. 2023 Mar 01. 14(1): 1172

Transgenic NADH dehydrogenase restores oxygen regulation of breathing in mitochondrial complex I-deficient mice.

Blanca Jiménez-Gómez, Patricia Ortega-Sáenz, Lin Gao, Patricia González-Rodríguez, Paula García-Flores, Navdeep Chandel, José López-Barneo.

The hypoxic ventilatory response (HVR) is a life-saving reflex, triggered by the activation of chemoreceptor glomus cells in the carotid body (CB) connected with the brainstem respiratory center. The molecular mechanisms underlying glomus cell acute oxygen (O2) sensing are unclear. Genetic disruption of mitochondrial complex I (MCI) selectively abolishes the HVR and glomus cell responsiveness to hypoxia. However, it is unknown what functions of MCI (metabolic, proton transport, or signaling) are essential for O2 sensing. Here we show that transgenic mitochondrial expression of NDI1, a single-molecule yeast NADH/quinone oxidoreductase that does not directly contribute to proton pumping, fully recovers the HVR and glomus cell sensitivity to hypoxia in MCI-deficient mice. Therefore, maintenance of mitochondrial NADH dehydrogenase activity and the electron transport chain are absolutely necessary for O2-dependent regulation of breathing. NDI1 expression also rescues other systemic defects caused by MCI deficiency. These data explain the role of MCI in acute O2 sensing by arterial chemoreceptors and demonstrate the optimal recovery of complex organismal functions by gene therapy.

DOI: https://doi.org/10.1038/s41467-023-36894-2
STAR Protoc. 2023 Jan 24. pii: S2666-1667(23)00022-9. [Epub ahead of print]4(1): 102064

Optimized protocol for quantification of mitochondrial non-heme and heme iron content in mouse tissues and cultured cells.

Tatsuya Sato, Hsiang-Chun Chang, Konrad T Sawicki, Hossein Ardehali.

  Impaired mitochondrial iron metabolism is associated with aging and a variety of diseases, and there is a growing need to accurately quantify mitochondrial iron levels. This protocol provides an optimized method for evaluating non-heme and heme iron in mitochondrial and cytosolic fractions of tissues and cultured cells. Our protocol consists of three steps: sample fractionation, non-heme iron measurement, and heme iron measurement. For complete details on the use and execution of this protocol, please refer to Sato et al. (2022).1.

Keywords:  Cell Biology; Cell Separation/Fractionation; Metabolism; Molecular Biology

DOI:  https://doi.org/10.1016/j.xpro.2023.102064
J Exp Bot. 2023 Feb 27. pii: erad062. [Epub ahead of print]

Assembly, transfer and fate of mitochondrial iron-sulfur clusters.

Luca Pedroletti, Anna Moseler, Andreas J Meyer.

  Since the discovery of an autonomous iron-sulfur cluster (Fe-S) assembly machinery in mitochondria, significant efforts to examine the nature of this process were made. The assembly of Fe-S clusters occurs in two distinct steps with the initial synthesis of [2Fe-2S] clusters by a first machinery followed by a subsequent assembly into [4Fe-4S] clusters by a second machinery. Despite this knowledge, we still have only a rudimentary understanding of how Fe-S clusters are transferred and distributed among their respective apoproteins. Especially considering that continuous protein turnover and particularly sacrificial destruction of clusters for synthesis of biotin and lipoic acid reveals possible bottlenecks in the supply chain of Fe-S clusters. Considering available information from other species, this review explores the mitochondrial assembly machinery of Arabidopsis and provides the current knowledge about the respective transfer steps to apoproteins. Furthermore, this review highlights biotin synthase and lipoyl synthase, which both utilize Fe-S clusters as sulfur source. After extraction of sulfur atoms from these clusters, the remains of the clusters likely fall apart releasing sulfide as a highly toxic by-product. Immediate refixation through local cysteine biosynthesis is therefore an essential salvage pathway and emphasises the physiological need for cysteine biosynthesis in plant mitochondria.

Keywords:  Biotin synthase; dehydrogenase complexes; glutaredoxin S15; iron-sulfur cluster; lipoic acid; lipoyl synthase; mitochondria; respiratory electron transport chain; sulfide

DOI:  https://doi.org/10.1093/jxb/erad062
Redox Biol. 2023 Feb 24. pii: S2213-2317(23)00044-7. [Epub ahead of print]61 102643

Mitochondrial cholesterol: Metabolism and impact on redox biology and disease.

Leire Goicoechea, Laura Conde de la Rosa, Sandra Torres, Carmen García-Ruiz, José C Fernández-Checa.

  Cholesterol is a crucial component of membrane bilayers by regulating their structural and functional properties. Cholesterol traffics to different cellular compartments including mitochondria, whose cholesterol content is low compared to other cell membranes. Despite the limited availability of cholesterol in the inner mitochondrial membrane (IMM), the metabolism of cholesterol in the IMM plays important physiological roles, acting as the precursor for the synthesis of steroid hormones and neurosteroids in steroidogenic tissues and specific neurons, respectively, or the synthesis of bile acids through an alternative pathway in the liver. Accumulation of cholesterol in mitochondria above physiological levels has a negative impact on mitochondrial function through several mechanisms, including the limitation of crucial antioxidant defenses, such as the glutathione redox cycle, increased generation of reactive oxygen species and consequent oxidative modification of cardiolipin, and defective assembly of respiratory supercomplexes. These adverse consequences of increased mitochondrial cholesterol trafficking trigger the onset of oxidative stress and cell death, and, ultimately, contribute to the development of diverse diseases, including metabolic liver diseases (i.e. fatty liver disease and liver cancer), as well as lysosomal disorders (i.e. Niemann-Pick type C disease) and neurodegenerative diseases (i.e. Alzheimer's disease). In this review, we summarize the metabolism and regulation of mitochondrial cholesterol and its potential impact on liver and neurodegenerative diseases.

Keywords:  Cholesterol; Liver disease; Metabolism; Mitochondria; Neurodegeneration; ROS

DOI:  https://doi.org/10.1016/j.redox.2023.102643
Indian J Radiol Imaging. 2023 Jan;33(1): 132-135

Leukodystrophy Associated with Mitochondrial Complex 1 Deficiency Due to Mutation in NUBPL Gene-An Unusual Follow-Up Finding.

Babu Peter S, Sree Vandana G.

DOI: https://doi.org/10.1055/s-0042-1758195
Parkinsonism Relat Disord. 2023 Feb 27. pii: S1353-8020(23)00076-7. [Epub ahead of print]109 105353

A novel C19orf12 frameshift mutation in a MPAN pedigree impairs mitochondrial function and connectivity leading to neurodegeneration.

Huan-Yun Chen, Han-I Lin, Chia-Lang Hsu, Pei-Lung Chen, Cheng-Yen Huang, Shu-Chun Teng, Chin-Hsien Lin.

BACKGROUND: Mitochondrial membrane protein‒associated neurodegeneration (MPAN) is a rare genetic disease characterized by progressive neurodegeneration with brain iron accumulations combined with neuronal α-synuclein and tau aggregations. Mutations in C19orf12 have been associated with both autosomal recessive and autosomal dominant inheritance patterns of MPAN.
METHODS: We present clinical features and functional evidence from a Taiwanese family with autosomal dominant MPAN caused by a novel heterozygous frameshift and nonsense mutation in C19orf12, c273_274 insA (p.P92Tfs*9). To verify the pathogenicity of the identified variant, we examined the mitochondrial function, morphology, protein aggregation, neuronal apoptosis, and RNA interactome in p.P92Tfs*9 mutant knock-in SH-SY5Y cells created with CRISPR-Cas9 technology.
RESULTS: Clinically, the patients with the C19orf12 p.P92Tfs*9 mutation presented with generalized dystonia, retrocollis, cerebellar ataxia, and cognitive decline, starting in their mid-20s. The identified novel frameshift mutation is located in the evolutionarily conserved region of the last exon of C19orf12. In vitro studies revealed that the p.P92Tfs*9 variant is associated with impaired mitochondrial function, reduced ATP production, aberrant mitochondria interconnectivity and ultrastructure. Increased neuronal α-synuclein and tau aggregations, and apoptosis were observed under conditions of mitochondrial stress. Transcriptomic analysis revealed that the expression of genes in clusters related to mitochondrial fission, lipid metabolism, and iron homeostasis pathways was altered in the C19orf12 p.P92Tfs*9 mutant cells compared to control cells.
CONCLUSION: Our findings provide clinical, genetic, and mechanistic insight revealing a novel heterozygous C19orf12 frameshift mutation to be a cause of autosomal dominant MPAN, further strengthening the importance of mitochondrial dysfunction in the pathogenesis of MPAN.

DOI: https://doi.org/10.1016/j.parkreldis.2023.105353
Cell Death Dis. 2023 Feb 27. 14(2): 162

Dysfunctional mitochondria accumulate in a skeletal muscle knockout model of Smn1, the causal gene of spinal muscular atrophy.

Francesco Chemello, Michela Pozzobon, Lorenza Iolanda Tsansizi, Tatiana Varanita, Rubèn Quintana-Cabrera, Daniele Bonesso, Martina Piccoli, Gerolamo Lanfranchi, Marta Giacomello, Luca Scorrano, Camilla Bean.

The approved gene therapies for spinal muscular atrophy (SMA), caused by loss of survival motor neuron 1 (SMN1), greatly ameliorate SMA natural history but are not curative. These therapies primarily target motor neurons, but SMN1 loss has detrimental effects beyond motor neurons and especially in muscle. Here we show that SMN loss in mouse skeletal muscle leads to accumulation of dysfunctional mitochondria. Expression profiling of single myofibers from a muscle specific Smn1 knockout mouse model revealed down-regulation of mitochondrial and lysosomal genes. Albeit levels of proteins that mark mitochondria for mitophagy were increased, morphologically deranged mitochondria with impaired complex I and IV activity and respiration and that produced excess reactive oxygen species accumulated in Smn1 knockout muscles, because of the lysosomal dysfunction highlighted by the transcriptional profiling. Amniotic fluid stem cells transplantation that corrects the SMN knockout mouse myopathic phenotype restored mitochondrial morphology and expression of mitochondrial genes. Thus, targeting muscle mitochondrial dysfunction in SMA may complement the current gene therapy.

DOI: https://doi.org/10.1038/s41419-023-05573-x
Viruses. 2023 Jan 26. pii: 351. [Epub ahead of print]15(2):

Mitochondria Dysfunction at the Heart of Viral Myocarditis: Mechanistic Insights and Therapeutic Implications.

Yasir Mohamud, Boaz Li, Amirhossein Bahreyni, Honglin Luo.

  The myocardium/heart is the most mitochondria-rich tissue in the human body with mitochondria comprising approximately 30% of total cardiomyocyte volume. As the resident "powerhouse" of cells, mitochondria help to fuel the high energy demands of a continuously beating myocardium. It is no surprise that mitochondrial dysfunction underscores the pathogenesis of many cardiovascular ailments, including those of viral origin such as virus-induced myocarditis. Enteroviruses have been especially linked to injuries of the myocardium and its sequelae dilated cardiomyopathy for which no effective therapies currently exist. Intriguingly, recent mechanistic insights have demonstrated viral infections to directly damage mitochondria, impair the mitochondrial quality control processes of the cell, such as disrupting mitochondrial antiviral innate immune signaling, and promoting mitochondrial-dependent pathological inflammation of the infected myocardium. In this review, we briefly highlight recent insights on the virus-mitochondria crosstalk and discuss the therapeutic implications of targeting mitochondria to preserve heart function and ultimately combat viral myocarditis.

Keywords:  apoptosis; autophagy; enterovirus; innate immune response; mitochondria; mitophagy; myocarditis

DOI:  https://doi.org/10.3390/v15020351
EMBO Rep. 2023 Feb 27. e54731

Saturated fatty acids increase LPI to reduce FUNDC1 dimerization and stability and mitochondrial function.

Linbo Chen, Qianping Zhang, Yuanyuan Meng, Tian Zhao, Chenglong Mu, Changying Fu, Caijuan Deng, Jianyu Feng, Siling Du, Wei Liu, Guangfeng Geng, Kaili Ma, Hongcheng Cheng, Qiangqiang Liu, Qian Luo, Jiaojiao Zhang, Zhanqiang Du, Lin Cao, Hui Wang, Yong Liu, Jianping Lin, Guo Chen, Lei Liu, Sin Man Lam, Guanghou Shui, Yushan Zhu, Quan Chen.

  Ectopic lipid deposition and mitochondrial dysfunction are common etiologies of obesity and metabolic disorders. Excessive dietary uptake of saturated fatty acids (SFAs) causes mitochondrial dysfunction and metabolic disorders, while unsaturated fatty acids (UFAs) counterbalance these detrimental effects. It remains elusive how SFAs and UFAs differentially signal toward mitochondria for mitochondrial performance. We report here that saturated dietary fatty acids such as palmitic acid (PA), but not unsaturated oleic acid (OA), increase lysophosphatidylinositol (LPI) production to impact on the stability of the mitophagy receptor FUNDC1 and on mitochondrial quality. Mechanistically, PA shifts FUNDC1 from dimer to monomer via enhanced production of LPI. Monomeric FUNDC1 shows increased acetylation at K104 due to dissociation of HDAC3 and increased interaction with Tip60. Acetylated FUNDC1 can be further ubiquitinated by MARCH5 for proteasomal degradation. Conversely, OA antagonizes PA-induced accumulation of LPI, and FUNDC1 monomerization and degradation. A fructose-, palmitate-, and cholesterol-enriched (FPC) diet also affects FUNDC1 dimerization and promotes its degradation in a non-alcoholic steatohepatitis (NASH) mouse model. We thus uncover a signaling pathway that orchestrates lipid metabolism with mitochondrial quality.

Keywords:  FUNDC1; fatty acid metabolism; membrane protein dimerization; mitochondrial quality control

DOI:  https://doi.org/10.15252/embr.202254731
Nat Rev Genet. 2023 Mar 02.

Methods and applications for single-cell and spatial multi-omics.

Katy Vandereyken, Alejandro Sifrim, Bernard Thienpont, Thierry Voet.

The joint analysis of the genome, epigenome, transcriptome, proteome and/or metabolome from single cells is transforming our understanding of cell biology in health and disease. In less than a decade, the field has seen tremendous technological revolutions that enable crucial new insights into the interplay between intracellular and intercellular molecular mechanisms that govern development, physiology and pathogenesis. In this Review, we highlight advances in the fast-developing field of single-cell and spatial multi-omics technologies (also known as multimodal omics approaches), and the computational strategies needed to integrate information across these molecular layers. We demonstrate their impact on fundamental cell biology and translational research, discuss current challenges and provide an outlook to the future.

DOI: https://doi.org/10.1038/s41576-023-00580-2
Front Pharmacol. 2023 ;14 1144093

Editorial: Mitochondrial therapy in neurodegeneration.

Anuradha Kalani, Pankaj Chaturvedi, Dario Brunetti, Komal Kalani.



Keywords:  Alzheimer’s; Parkinson’s; mitochondria; neurodegeneration; therapy

DOI:  https://doi.org/10.3389/fphar.2023.1144093
Hum Genomics. 2023 02 27. 17(1): 14

Computational and mitochondrial functional studies of novel compound heterozygous variants in SPATA5 gene support a causal link with epileptogenic encephalopathy.

Víctor Raggio, Martín Graña, Erik Winiarski, Santiago Mansilla, Camila Simoes, Soledad Rodríguez, Mariana Brandes, Alejandra Tapié, Laura Rodríguez, Lucía Cibils, Martina Alonso, Jennyfer Martínez, Tamara Fernández-Calero, Fernanda Domínguez, Melania Rosas Mezquida, Laura Castro, Alfredo Cerisola, Hugo Naya, Adriana Cassina, Celia Quijano, Lucía Spangenberg.

The SPATA5 gene encodes a 892 amino-acids long protein that has a putative mitochondrial targeting sequence and has been proposed to function in maintenance of mitochondrial function and integrity during mouse spermatogenesis. Several studies have associated homozygous or compound heterozygous mutations in SPATA5 gene to microcephaly, intellectual disability, seizures and hearing loss. This suggests a role of the SPATA5 gene also in neuronal development. Recently, our group presented results validating the use of blood cells for the assessment of mitochondrial function for diagnosis and follow-up of mitochondrial disease, minimizing the need for invasive procedures such as muscle biopsy. In this study, we were able to diagnose a patient with epileptogenic encephalopathy using next generation sequencing. We found two novel compound heterozygous variants in SPATA5 that are most likely causative. To analyze the impact of SPATA5 mutations on mitochondrial functional studies directly on the patients' mononuclear cells and platelets were undertaken. Oxygen consumption rates in platelets and PBMCs were impaired in the patient when compared to a healthy control. Also, a decrease in mitochondrial mass was observed in the patient monocytes with respect to the control. This suggests a true pathogenic effect of the mutations in mitochondrial function, especially in energy production and possibly biogenesis, leading to the observed phenotype.

DOI: https://doi.org/10.1186/s40246-023-00463-x
Cell Mol Neurobiol. 2023 Mar 03.

Rescuers from the Other Shore: Intercellular Mitochondrial Transfer and Its Implications in Central Nervous System Injury and Diseases.

Weichen Dong, Wenxin Zhang, Linying Yuan, Yi Xie, Yunzi Li, Kuanyu Li, Wusheng Zhu.

  As the powerhouse and core of cellular metabolism and survival, mitochondria are the essential organelle in mammalian cells and maintain cellular homeostasis by changing their content and morphology to meet demands through mitochondrial quality control. It has been observed that mitochondria can move between cells under physiological and pathophysiological conditions, which provides a novel strategy for preserving mitochondrial homeostasis and also a therapeutic target for applications in clinical settings. Therefore, in this review, we will summarize currently known mechanisms of intercellular mitochondrial transfer, including modes, triggers, and functions. Due to the highly demanded energy and indispensable intercellular linkages of the central nervous system (CNS), we highlight the mitochondrial transfer in CNS. We also discuss future application possibilities and difficulties that need to be addressed in the treatment of CNS injury and diseases. This clarification should shed light on its potential clinical applications as a promising therapeutic target in neurological diseases. Intercellular mitochondrial transfer maintains the homeostasis of central nervous system (CNS), and its alteration is related to several neurological diseases. Supplementing exogenous mitochondrial donor cells and mitochondria, or utilizing some medications to regulate the process of transfer might mitigate the disease and injury.

Keywords:  Central nervous system; Intercellular mitochondrial transfer; Mitochondria; Mitochondrial quality control; Mitochondrial transplantation

DOI:  https://doi.org/10.1007/s10571-023-01331-x
Exp Cell Res. 2023 Feb 27. pii: S0014-4827(23)00083-6. [Epub ahead of print]425(1): 113536

Inhibition of mitochondrial transcription by the neurotoxin MPP.

Alexander Pfab, Sergey Belikov, Michaela Keuper, Martin Jastroch, Mattias Mannervik.

  The neurotoxin MPP+ triggers cell death of dopamine neurons and induces Parkinson's disease symptoms in mice and men, but the immediate transcriptional response to this neurotoxin has not been studied. We therefore treated human SH-SY5Y cells with a low dose (0.1 mM) of MPP+ and measured the effect on nascent transcription by precision run-on sequencing (PRO-seq). We found that transcription of the mitochondrial genome was significantly reduced already after 30 min, whereas nuclear gene transcription was unaffected. Inhibition of respiratory complex I by MPP+ led to reduced ATP production, that may explain the diminished activity of mitochondrial RNA polymerase. Our results show that MPP+ has a direct effect on mitochondrial function and transcription, and that other gene expression or epigenetic changes induced by this neurotoxin are secondary effects that reflect a cellular adaptation program.

Keywords:  ATP; Mitochondria; Neurotoxin; Parkinson's disease; Respiration; Transcription

DOI:  https://doi.org/10.1016/j.yexcr.2023.113536
Orphanet J Rare Dis. 2023 Mar 02. 18(1): 43

Mitochondrial diseases in Hong Kong: prevalence, clinical characteristics and genetic landscape.

Tsz-Sum Wong, Kiran M Belaramani, Chun-Kong Chan, Wing-Ki Chan, Wai-Lun Larry Chan, Shek-Kwan Chang, Sing-Ngai Cheung, Ka-Yin Cheung, Yuk-Fai Cheung, Shuk-Ching Josephine Chong, Chi-Kwan Jasmine Chow, Hon-Yin Brian Chung, Sin-Ying Florence Fan, Wai-Ming Joshua Fok, Ka-Wing Fong, Tsui-Hang Sharon Fung, Kwok-Fai Hui, Ting-Hin Hui, Joannie Hui, Chun-Hung Ko, Min-Chung Kwan, Mei-Kwan Anne Kwok, Sung-Shing Jeffrey Kwok, Moon-Sing Lai, Yau-On Lam, Ching-Wan Lam, Ming-Chung Lau, Chun-Yiu Eric Law, Wing-Cheong Lee, Han-Chih Hencher Lee, Chin-Nam Lee, Kin-Hang Leung, Kit-Yan Leung, Siu-Hung Li, Tsz-Ki Jacky Ling, Kam-Tim Timothy Liu, Fai-Man Lo, Hiu-Tung Lui, Ching-On Luk, Ho-Ming Luk, Che-Kwan Ma, Karen Ma, Kam-Hung Ma, Yuen-Ni Mew, Alex Mo, Sui-Fun Ng, Wing-Kit Grace Poon, Richard Rodenburg, Bun Sheng, Jan Smeitink, Cheuk-Ling Charing Szeto, Shuk-Mui Tai, Choi-Ting Alan Tse, Li-Yan Lilian Tsung, Ho-Ming June Wong, Wing-Yin Winnie Wong, Kwok-Kui Wong, Suet-Na Sheila Wong, Chun-Nei Virginia Wong, Wai-Shan Sammy Wong, Chi-Kin Felix Wong, Shun-Ping Wu, Hiu-Fung Jerome Wu, Man-Mut Yau, Kin-Cheong Eric Yau, Wai-Lan Yeung, Hon-Ming Jonas Yeung, Kin-Keung Edwin Yip, Pui-Hong Terence Young, Gao Yuan, Yuet-Ping Liz Yuen, Chi-Lap Yuen, Cheuk-Wing Fung.

   OBJECTIVE: To determine the prevalence of mitochondrial diseases (MD) in Hong Kong (HK) and to evaluate the clinical characteristics and genetic landscape of MD patients in the region.
METHODS: This study retrospectively reviewed the phenotypic and molecular characteristics of MD patients from participating public hospitals in HK between January 1985 to October 2020. Molecularly and/or enzymatically confirmed MD cases of any age were recruited via the Clinical Analysis and Reporting System (CDARS) using relevant keywords and/or International Classification of Disease (ICD) codes under the HK Hospital Authority or through the personal recollection of treating clinicians among the investigators.
RESULTS: A total of 119 MD patients were recruited and analyzed in the study. The point prevalence of MD in HK was 1.02 in 100,000 people (95% confidence interval 0.81-1.28 in 100,000). 110 patients had molecularly proven MD and the other nine were diagnosed by OXPHOS enzymology analysis or mitochondrial DNA depletion analysis with unknown molecular basis. Pathogenic variants in the mitochondrial genome (72 patients) were more prevalent than those in the nuclear genome (38 patients) in our cohort. The most commonly involved organ system at disease onset was the neurological system, in which developmental delay, seizures or epilepsy, and stroke-like episodes were the most frequently reported presentations. The mortality rate in our cohort was 37%.
CONCLUSION: This study is a territory-wide overview of the clinical and genetic characteristics of MD patients in a Chinese population, providing the first available prevalence rate of MD in Hong Kong. The findings of this study aim to facilitate future in-depth evaluation of MD and lay the foundation to establish a local MD registry.

Keywords:  Hong Kong; Mitochondrial diseases; Prevalence

DOI:  https://doi.org/10.1186/s13023-023-02632-6
Aging (Albany NY). 2023 Feb 27. 15

Aging and memory are altered by genetically manipulating lactate dehydrogenase in the neurons or glia of flies.

Ariel K Frame, J Wesley Robinson, Nader H Mahmoudzadeh, Jason M Tennessen, Anne F Simon, Robert C Cumming.

  The astrocyte-neuron lactate shuttle hypothesis posits that glial-generated lactate is transported to neurons to fuel metabolic processes required for long-term memory. Although studies in vertebrates have revealed that lactate shuttling is important for cognitive function, it is uncertain if this form of metabolic coupling is conserved in invertebrates or is influenced by age. Lactate dehydrogenase (Ldh) is a rate limiting enzyme that interconverts lactate and pyruvate. Here we genetically manipulated expression of Drosophila melanogaster lactate dehydrogenase (dLdh) in neurons or glia to assess the impact of altered lactate metabolism on invertebrate aging and long-term courtship memory at different ages. We also assessed survival, negative geotaxis, brain neutral lipids (the core component of lipid droplets) and brain metabolites. Both upregulation and downregulation of dLdh in neurons resulted in decreased survival and memory impairment with age. Glial downregulation of dLdh expression caused age-related memory impairment without altering survival, while upregulated glial dLdh expression lowered survival without disrupting memory. Both neuronal and glial dLdh upregulation increased neutral lipid accumulation. We provide evidence that altered lactate metabolism with age affects the tricarboxylic acid (TCA) cycle, 2-hydroxyglutarate (2HG), and neutral lipid accumulation. Collectively, our findings indicate that the direct alteration of lactate metabolism in either glia or neurons affects memory and survival but only in an age-dependent manner.

Keywords:  Drosophila melanogaster; astrocyte-neuron lactate shuttle (ANLS); courtship conditioning; dLdh; glia; lactate; lactate dehydrogenase; long-term memory

DOI:  https://doi.org/10.18632/aging.204565
Nat Commun. 2023 Mar 03. 14(1): 1230

RNA splicing analysis using heterogeneous and large RNA-seq datasets.

Jorge Vaquero-Garcia, Joseph K Aicher, San Jewell, Matthew R Gazzara, Caleb M Radens, Anupama Jha, Scott S Norton, Nicholas F Lahens, Gregory R Grant, Yoseph Barash.

The ubiquity of RNA-seq has led to many methods that use RNA-seq data to analyze variations in RNA splicing. However, available methods are not well suited for handling heterogeneous and large datasets. Such datasets scale to thousands of samples across dozens of experimental conditions, exhibit increased variability compared to biological replicates, and involve thousands of unannotated splice variants resulting in increased transcriptome complexity. We describe here a suite of algorithms and tools implemented in the MAJIQ v2 package to address challenges in detection, quantification, and visualization of splicing variations from such datasets. Using both large scale synthetic data and GTEx v8 as benchmark datasets, we assess the advantages of MAJIQ v2 compared to existing methods. We then apply MAJIQ v2 package to analyze differential splicing across 2,335 samples from 13 brain subregions, demonstrating its ability to offer insights into brain subregion-specific splicing regulation.

DOI: https://doi.org/10.1038/s41467-023-36585-y