bims-mitdis Biomed News
on Mitochondrial disorders
Issue of 2021‒09‒12
37 papers selected by
Catalina Vasilescu
University of Helsinki

  1. Hum Mol Genet. 2021 Sep 11. pii: ddab257. [Epub ahead of print]
      TARS2 encodes human mitochondrial threonyl tRNA-synthetase that is responsible for generating mitochondrial Thr-tRNAThr and clearing mischarged Ser-tRNAThr during mitochondrial translation. Pathogenic variants in TARS2 have hitherto been reported in a pair of siblings and an unrelated patient with an early onset mitochondrial encephalomyopathy and a combined respiratory chain enzyme deficiency in muscle. We here report five additional unrelated patients with TARS2-related mitochondrial diseases, expanding the clinical phenotype to also include epilepsy, dystonia, hyperhidrosis and severe hearing impairment. Additionally, we document seven novel TARS2 variants-one nonsense variant and six missense variants-that we demonstrate are pathogenic and causal of the disease presentation based on population frequency, homology modelling and functional studies that show the effects of the pathogenic variants on TARS2 stability and/or function.
  2. Autophagy. 2021 Sep 05. 1-3
      Among other mechanisms, mitochondrial membrane dynamics including mitochondrial fission and fusion, and the activity of the ubiquitin (Ub)-proteasome system (UPS) both are critical for maintaining mitochondrial function. To advance our knowledge of the role of mitochondrial fission, the UPS, and how they coordinatively affect mitochondrial response to proteotoxicity, we analyzed mitochondrial ubiquitination and mitochondria-specific autophagy (mitophagy) in E3 Ub ligase PRKN/parkin-expressing and -deficient cells. Through imaging, biochemical, and genetic analyses, we found that in a model of acute reduction of mitochondrial translation fidelity (MTF) some population of mitochondria within a single cell are enriched, while some showed reduced levels of CYCS (cytochrome c, somatic) and CPOX (coproporphyrinogen oxidase) proteins, both located in the intermembrane space (IMS); henceforth called "mosaic distribution". Formation of mosaic mitochondria requires mitochondrial fission and active mitochondrial translation. In cell lines deficient in PRKN activity, this process is followed by severing the outer mitochondrial membrane (OMM) and ubiquitination of the inner mitochondrial membrane (IMM) proteins (including TRAP1 and CPOX), recruitment of autophagy receptors, and formation of mito-autophagosomes. In contrast, in PRKN-expressing cells, mitochondria with high CYCS and CPOX levels are preferentially targeted by PRKN, leading to OMM ubiquitination and canonical PRKN-PINK1-mediated autophagy.
    Keywords:  DRP1; Parkin; mitochondria; mitochondrial translation; mitophagy; ubiquitin
  3. Nat Metab. 2021 Sep 09.
      Mitochondria are the main site for generating reactive oxygen species, which are key players in diverse biological processes. However, the molecular pathways of redox signal transduction from the matrix to the cytosol are poorly defined. Here we report an inside-out redox signal of mitochondria. Cysteine oxidation of MIC60, an inner mitochondrial membrane protein, triggers the formation of disulfide bonds and the physical association of MIC60 with Miro, an outer mitochondrial membrane protein. The oxidative structural change of this membrane-crossing complex ultimately elicits cellular responses that delay mitophagy, impair cellular respiration and cause oxidative stress. Blocking the MIC60-Miro interaction or reducing either protein, genetically or pharmacologically, extends lifespan and health-span of healthy fruit flies, and benefits multiple models of Parkinson's disease and Friedreich's ataxia. Our discovery provides a molecular basis for common treatment strategies against oxidative stress.
  4. Nat Struct Mol Biol. 2021 Sep 06.
      Human mitochondrial transcripts contain messenger and ribosomal RNAs flanked by transfer RNAs (tRNAs), which are excised by mitochondrial RNase (mtRNase) P and Z to liberate all RNA species. In contrast to nuclear or bacterial RNase P, mtRNase P is not a ribozyme but comprises three protein subunits that carry out RNA cleavage and methylation by unknown mechanisms. Here, we present the cryo-EM structure of human mtRNase P bound to precursor tRNA, which reveals a unique mechanism of substrate recognition and processing. Subunits TRMT10C and SDR5C1 form a subcomplex that binds conserved mitochondrial tRNA elements, including the anticodon loop, and positions the tRNA for methylation. The endonuclease PRORP is recruited and activated through interactions with its PPR and nuclease domains to ensure precise pre-tRNA cleavage. The structure provides the molecular basis for the first step of RNA processing in human mitochondria.
  5. J Biol Chem. 2021 Sep 01. pii: S0021-9258(21)00957-1. [Epub ahead of print] 101155
      Acylation modifications, such as the succinylation of lysine, are post-translational modifications and a powerful means of regulating protein activity. Some acylations occur nonenzymatically, driven by an increase in the concentration of acyl group donors. Lysine succinylation has a profound effect on the corresponding site within the protein, as it dramatically changes the charge of the residue. In eukaryotes, it predominantly affects mitochondrial proteins because the donor of succinate, succinyl-coenzyme A, is primarily generated in the tricarboxylic acid (TCA) cycle. Although numerous succinylated mitochondrial proteins have been identified in Saccharomyces cerevisiae, a more detailed characterization of the yeast mitochondrial succinylome is still lacking. Here we performed a proteomic mass spectrometry analysis of purified yeast mitochondria and detected 314 succinylated mitochondrial proteins with 1763 novel succinylation sites. The mitochondrial nucleoid, a complex of mitochondrial DNA (mtDNA) and mitochondrial proteins, is one of the structures whose protein components are affected by succinylation. We found that Abf2p, the principal component of mt-nucleoids responsible for compacting mtDNA in S. cerevisiae, can be succinylated in vivo on at least thirteen lysine residues. Abf2p succinylation in vitro inhibits its DNA-binding activity and reduces its sensitivity to digestion by the ATP-dependent ScLon protease. We conclude that changes in the metabolic state of a cell resulting in an increase in concentration of TCA intermediates may affect mitochondrial functions.
    Keywords:  DNA-protein interaction; lysine succinylation; mitochondria; mitochondrial DNA; mitochondrial nucleoid; post-translational modification (PTM); proteomics; succinylome; yeast
  6. Mol Biol Cell. 2021 Sep 08. mbcE21040191
      Mitochondria evolved from endosymbiotic bacteria to become essential organelles of eukaryotic cells. The unique lipid composition and structure of mitochondrial membranes are critical for the proper functioning of mitochondria. However, stress responses that help maintain the mitochondrial membrane integrity are not well understood. One reason for this lack of insight is the absence of efficient tools to specifically damage mitochondrial membranes. Here, through a compound screen, we found that two bis-biguanide compounds, Chlorhexidine and Alexidine, modified the activity of the inner mitochondrial membrane (IMM)-resident protease OMA1 by altering the integrity of the IMM. These compounds are well-known bactericides whose mechanism of action has centered on their damage-inducing activity on bacterial membranes. We found Alexidine binds to the IMM likely through the electrostatic interaction driven by the membrane potential as well as an affinity for anionic phospholipids. Electron microscopic analysis revealed that Alexidine severely perturbated the cristae structure. Notably, Alexidine evoked a specific transcriptional/proteostasis signature that was not induced by other typical mitochondrial stressors, highlighting the unique property of Alexidine as a novel mitochondrial membrane stressor. Our findings provide a chemical-biological tool that should enable the delineation of mitochondrial stress-signaling pathways required to maintain the mitochondrial membrane homeostasis.
  7. Int J Mol Sci. 2021 Sep 06. pii: 9655. [Epub ahead of print]22(17):
      Most mitochondrial proteins are synthesized in the cytosol and targeted to the mitochondrial surface in a post-translational manner. The surface of the endoplasmic reticulum (ER) plays an active role in this targeting reaction. ER-associated chaperones interact with certain mitochondrial membrane protein precursors and transfer them onto receptor proteins of the mitochondrial surface in a process termed ER-SURF. ATP-driven proteins in the membranes of mitochondria (Msp1, ATAD1) and the ER (Spf1, P5A-ATPase) serve as extractors for the removal of mislocalized proteins. If the re-routing to mitochondria fails, precursors can be degraded by ER or mitochondria-associated degradation (ERAD or MAD respectively) in a proteasome-mediated reaction. This review summarizes the current knowledge about the cooperation of the ER and mitochondria in the targeting and quality control of mitochondrial precursor proteins.
    Keywords:  ER-SURF; chaperones; contact sites; endoplasmic reticulum; membrane extraction; mitochondria; protein targeting
  8. Wiley Interdiscip Rev RNA. 2021 Sep 08. e1690
      Mitochondria play a pivotal role in numerous cellular processes. One of them is regulation of the innate immune pathway. In this instance, mitochondria function in two different aspects of regulatory mechanisms. First, mitochondria are part of the antiviral signaling cascade that is triggered in the cytoplasm and transmitted to effector proteins through mitochondria-localized proteins. Second, mitochondria can become an endogenous source of innate immune stimuli. Under some pathophysiological conditions, mitochondria release to the cytoplasm immunogenic factors, such as mitochondrial nucleic acids. Here, we focus on immunogenic mitochondrial double-stranded RNA (mt-dsRNA) and its origin and metabolism. We discuss factors that are responsible for regulating mt-dsRNA and its escape from mitochondria, emphasizing the contribution of polynucleotide phosphorylase (PNPase, PNPT1). Finally, we review current knowledge of the role of PNPase in human health and disease. This article is categorized under: RNA in Disease and Development > RNA in Disease.
    Keywords:  innate immunity; mitochondrial RNA decay and surveillance; mitochondrial dsRNA
  9. Int J Mol Sci. 2021 Sep 01. pii: 9502. [Epub ahead of print]22(17):
      The human mitochondrial genome (mtDNA) regulates its transcription products in specialised and distinct ways as compared to nuclear transcription. Thanks to its mtDNA mitochondria possess their own set of tRNAs, rRNAs and mRNAs that encode a subset of the protein subunits of the electron transport chain complexes. The RNA regulation within mitochondria is organised within specialised, membraneless, compartments of RNA-protein complexes, called the Mitochondrial RNA Granules (MRGs). MRGs were first identified to contain nascent mRNA, complexed with many proteins involved in RNA processing and maturation and ribosome assembly. Most recently, double-stranded RNA (dsRNA) species, a hybrid of the two complementary mRNA strands, were found to form granules in the matrix of mitochondria. These RNA granules are therefore components of the mitochondrial post-transcriptional pathway and as such play an essential role in mitochondrial gene expression. Mitochondrial dysfunctions in the form of, for example, RNA processing or RNA quality control defects, or inhibition of mitochondrial fission, can cause the loss or the aberrant accumulation of these RNA granules. These findings underline the important link between mitochondrial maintenance and the efficient expression of its genome.
    Keywords:  RNA degradation; RNA processing; degradosome; dsRNA; liquid–liquid phase separation (LLPS); mitochondrial RNA granules (MRGs); mitochondrial gene expression; nucleoids
  10. Front Genet. 2021 ;12 725259
      Genetic disorders are a frequent cause of hospitalization, morbidity and mortality in pediatric patients, especially in the neonatal or pediatric intensive care unit (NICU/PICU). In recent years, rapid genome-wide sequencing (exome or whole genome sequencing) has been applied in the NICU/PICU. However, mtDNA sequencing is not routinely available in rapid genetic diagnosis programs, which may fail to diagnose mtDNA mutation-associated diseases. Herein, we explored the clinical utility of rapid exome sequencing combined with mtDNA sequencing in critically ill pediatric patients with suspected genetic disorders. Rapid clinical exome sequencing (CES) was performed as a first-tier test in 40 critically ill pediatric patients (aged from 6 days to 15 years) with suspected genetic conditions. Blood samples were also collected from the parents for trio analysis. Twenty-six patients presented with neuromuscular abnormalities or other systemic abnormalities, suggestive of suspected mitochondrial diseases or the necessity for a differential diagnosis of other diseases, underwent rapid mtDNA sequencing concurrently. A diagnosis was made in 18 patients (45.0%, 18/40); three cases with de novo autosomal dominant variants, ten cases with homozygous or compound heterozygous variants, three cases with hemizygous variants inherited from mother, three cases with heterozygous variants inherited from either parent, and one case with a mtDNA mutation. The 18 patients were diagnosed with metabolic (n = 7), immunodeficiency (n = 4), cardiovascular (n = 2), neuromuscular (n = 2) disorders, and others. Genetic testing reports were generated with a median time of 5 days (range, 3-9 days). Thirteen patients that were diagnosed had an available medical treatment and resulted in a positive outcome. We propose that rapid exome sequencing combined with mitochondrial DNA sequencing should be available to patients with suspected mitochondrial diseases or undefined clinical features necessary for making a differential diagnosis of other diseases.
    Keywords:  genetic disorders; mitochondrial diseases; mtDNA sequencing; pediatric patients; rapid exome sequencing
  11. Neuron. 2021 Sep 01. pii: S0896-6273(21)00608-5. [Epub ahead of print]
      Neurons require mechanisms to maintain ATP homeostasis in axons, which are highly vulnerable to bioenergetic failure. Here, we elucidate a transcellular signaling mechanism by which oligodendrocytes support axonal energy metabolism via transcellular delivery of NAD-dependent deacetylase SIRT2. SIRT2 is undetectable in neurons but enriched in oligodendrocytes and released within exosomes. By deleting sirt2, knocking down SIRT2, or blocking exosome release, we demonstrate that transcellular delivery of SIRT2 is critical for axonal energy enhancement. Mass spectrometry and acetylation analyses indicate that neurons treated with oligodendrocyte-conditioned media from WT, but not sirt2-knockout, mice exhibit strong deacetylation of mitochondrial adenine nucleotide translocases 1 and 2 (ANT1/2). In vivo delivery of SIRT2-filled exosomes into myelinated axons rescues mitochondrial integrity in sirt2-knockout mouse spinal cords. Thus, our study reveals an oligodendrocyte-to-axon delivery of SIRT2, which enhances ATP production by deacetylating mitochondrial proteins, providing a target for boosting axonal bioenergetic metabolism in neurological disorders.
    Keywords:  acetylation; adenine nucleotide translocases 1 and 2; axonal ATP; axonal energetics; axonal mitochondria; energy metabolism; exosome; myelin; oligodendrocyte; sirtuin 2
  12. Nat Commun. 2021 Sep 09. 12(1): 5354
      Mitochondrial division is not an autonomous event but involves multiple organelles, including the endoplasmic reticulum (ER) and lysosomes. Whereas the ER drives the constriction of mitochondrial membranes, the role of lysosomes in mitochondrial division is not known. Here, using super-resolution live-cell imaging, we investigate the recruitment of lysosomes to the site of mitochondrial division. We find that the ER recruits lysosomes to the site of division through the interaction of VAMP-associated proteins (VAPs) with the lysosomal lipid transfer protein ORP1L to induce a three-way contact between the ER, lysosome, and the mitochondrion. We also show that ORP1L might transport phosphatidylinositol-4-phosphate (PI(4)P) from lysosomes to mitochondria, as inhibiting its transfer or depleting PI(4)P at the mitochondrial division site impairs fission, demonstrating a direct role for PI(4)P in the division process. Our findings support a model where the ER recruits lysosomes to act in concert at the fission site for the efficient division of mitochondria.
  13. Hum Mol Genet. 2021 Sep 07. pii: ddab254. [Epub ahead of print]
      Calcium signaling via mitochondrial calcium uniporter (MCU) complex coordinates mitochondrial bioenergetics with cellular energy demands. Emerging studies show that the stability and activity of the pore-forming subunit of the complex, MCU, is dependent on the mitochondrial phospholipid, cardiolipin (CL), but how this impacts calcium-dependent mitochondrial bioenergetics in CL-deficiency disorder like Barth syndrome (BTHS) is not known. Here we utilized multiple models of BTHS including yeast, mouse muscle cell line, as well as BTHS patient cells and cardiac tissue to show that CL is required for the abundance and stability of the MCU-complex regulatory subunit MICU1. Interestingly, the reduction in MICU1 abundance in BTHS mitochondria is independent of MCU. Unlike MCU and MICU1/MICU2, other subunit and associated factor of the uniporter complex, EMRE and MCUR1, respectively, are not affected in BTHS models. Consistent with the decrease in MICU1 levels, we show that the kinetics of MICU1-dependent mitochondrial calcium uptake is perturbed and acute stimulation of mitochondrial calcium signaling in BTHS myoblasts fails to activate pyruvate dehydrogenase, which in turn impairs the generation of reducing equivalents and blunts mitochondrial bioenergetics. Taken together, our findings suggest that defects in mitochondrial calcium signaling could contribute to cardiac and skeletal muscle pathologies observed in BTHS patients.
  14. JCI Insight. 2021 Sep 08. pii: 138088. [Epub ahead of print]6(17):
      Mitochondrial dysfunction is a major pathophysiological contributor to the progression of Parkinson's disease (PD); however, whether it contributes to epigenetic dysregulation remains unknown. Here, we show that both chemically and genetically driven mitochondrial dysfunctions share a common mechanism of epigenetic dysregulation. Under both scenarios, lysine 27 acetylation of likely variant H3.3 (H3.3K27ac) increased in dopaminergic neuronal models of PD, thereby opening that region to active enhancer activity via H3K27ac. These vulnerable epigenomic loci represent potential transcription factor motifs for PD pathogenesis. We further confirmed that mitochondrial dysfunction induces H3K27ac in ex vivo and in vivo (MitoPark) neurodegenerative models of PD. Notably, the significantly increased H3K27ac in postmortem PD brains highlights the clinical relevance to the human PD population. Our results reveal an exciting mitochondrial dysfunction-metabolism-H3K27ac-transcriptome axis for PD pathogenesis. Collectively, the mechanistic insights link mitochondrial dysfunction to epigenetic dysregulation in dopaminergic degeneration and offer potential new epigenetic intervention strategies for PD.
    Keywords:  Epigenetics; Mitochondria; Neuroscience; Parkinson disease
  15. Elife. 2021 09 07. pii: e62091. [Epub ahead of print]10
      Developing neurons form synapses at a high rate. Synaptic transmission is very energy-demanding and likely requires ATP production by mitochondria nearby. Mitochondria might be targeted to active synapses in young dendrites, but whether such motility regulation mechanisms exist is unclear. We investigated the relationship between mitochondrial motility and neuronal activity in the primary visual cortex of young mice in vivo and in slice cultures. During the first 2 postnatal weeks, mitochondrial motility decreases while the frequency of neuronal activity increases. Global calcium transients do not affect mitochondrial motility. However, individual synaptic transmission events precede local mitochondrial arrest. Pharmacological stimulation of synaptic vesicle release, but not focal glutamate application alone, stops mitochondria, suggesting that an unidentified factor co-released with glutamate is required for mitochondrial arrest. A computational model of synaptic transmission-mediated mitochondrial arrest shows that the developmental increase in synapse number and transmission frequency can contribute substantially to the age-dependent decrease of mitochondrial motility.
    Keywords:  calcium signaling; in vivo imaging; intracellular transport; mouse; neuroscience; synaptic transmission
  16. Intern Med J. 2021 Sep 09.
      This document provides consensus-based recommendations for general physicians and primary care physicians who diagnose and manage patients with mitochondrial diseases (MDs). It builds on previous international guidelines,1 with particular emphasis on clinical management in the Australian setting. This statement was prepared by a working group of medical practitioners, nurses and allied health professionals with clinical expertise and experience in managing Australian patients with MD. As new treatments and management plans emerge, these consensus-based recommendations will continue to evolve, but current standards of care are summarised in this document. This article is protected by copyright. All rights reserved.
    Keywords:  clinical guidelines; mitochondrial disease; treatment
  17. Mitochondrion. 2021 Sep 06. pii: S1567-7249(21)00118-5. [Epub ahead of print]
      Topoisomerases regulate DNA topology, organization of the intracellular DNA, the transmission of genetic materials, and gene expressions. Other than the nuclear genome, mitochondria also harbor the small, circular DNA (mtDNA) that encodes a critical subset of proteins for the production of cellular ATP; however, mitochondria are solely dependent on the nucleus for all the mitochondrial proteins necessary for mtDNA replication, repair, and maintenance. Mitochondrial genome compiles topological stress from bidirectional transcription and replication, therefore imports four nuclear encoded topoisomerases (Top1mt, Top2α, Top2β, and Top3α) in the mitochondria to relax mtDNA supercoiling generated during these processes. Trapping of topoisomerase on DNA results in the formation of protein-linked DNA adducts (PDAs), which are widely exploited by topoisomerase-targeting anticancer drugs. Intriguingly mtDNA is potentially exposed to DNA damage that has been attributed to a variety of human diseases, including neurodegeneration, cancer, and premature aging. In this review, we focus on the role of different topoisomerases in the mitochondria and our current understanding of the mitochondrial DNA damage through trapped protein-DNA complexes, and the progress in the molecular mechanisms of the repair for trapped topoisomerase covalent complexes (Topcc). Finally, we have discussed how the pathological DNA lesions that cause mtDNA damage,trigger mitochondrial fission and mitophagy, which serve as quality control events for clearing damaged mtDNA.
    Keywords:  DNA repair; Mitochondria; TDP1; TDP2; TFAM; Topoisomerase 1; Topoisomerase II; mitochondrial DNA; neurological diseases
  18. Aging Cell. 2021 Sep 09. e13472
      Metabolic dysfunction and protein aggregation are common characteristics that occur in age-related neurodegenerative disease. However, the mechanisms underlying these abnormalities remain poorly understood. We have found that mutations in the gene encoding presenilin in Caenorhabditis elegans, sel-12, results in elevated mitochondrial activity that drives oxidative stress and neuronal dysfunction. Mutations in the human presenilin genes are the primary cause of familial Alzheimer's disease. Here, we demonstrate that loss of SEL-12/presenilin results in the hyperactivation of the mTORC1 pathway. This hyperactivation is caused by elevated mitochondrial calcium influx and, likely, the associated increase in mitochondrial activity. Reducing mTORC1 activity improves proteostasis defects and neurodegenerative phenotypes associated with loss of SEL-12 function. Consistent with high mTORC1 activity, we find that SEL-12 loss reduces autophagosome formation, and this reduction is prevented by limiting mitochondrial calcium uptake. Moreover, the improvements of proteostasis and neuronal defects in sel-12 mutants due to mTORC1 inhibition require the induction of autophagy. These results indicate that mTORC1 hyperactivation exacerbates the defects in proteostasis and neuronal function in sel-12 mutants and demonstrate a critical role of presenilin in promoting neuronal health.
    Keywords:   Caenorhabditis elegans ; Alzheimer; aging; calcium; mTORC1; mitochondria; presenilin
  19. Nucleic Acids Res. 2021 Sep 09. pii: gkab770. [Epub ahead of print]
      We report a rapid experimental procedure based on high-density in vivo psoralen inter-strand DNA cross-linking coupled to spreading of naked purified DNA, positive staining, low-angle rotary shadowing, and transmission electron microscopy (TEM) that allows quick visualization of the dynamic of heavy strand (HS) and light strand (LS) human mitochondrial DNA replication. Replication maps built on linearized mitochondrial genomes and optimized rotary shadowing conditions enable clear visualization of the progression of the mitochondrial DNA synthesis and visualization of replication intermediates carrying long single-strand DNA stretches. One variant of this technique, called denaturing spreading, allowed the inspection of the fine chromatin structure of the mitochondrial genome and was applied to visualize the in vivo three-strand DNA structure of the human mitochondrial D-loop intermediate with unprecedented clarity.
  20. Proc Natl Acad Sci U S A. 2021 Sep 14. pii: e2025932118. [Epub ahead of print]118(37):
      Mitochondria form a complex, interconnected reticulum that is maintained through coordination among biogenesis, dynamic fission, and fusion and mitophagy, which are initiated in response to various cues to maintain energetic homeostasis. These cellular events, which make up mitochondrial quality control, act with remarkable spatial precision, but what governs such spatial specificity is poorly understood. Herein, we demonstrate that specific isoforms of the cellular bioenergetic sensor, 5' AMP-activated protein kinase (AMPKα1/α2/β2/γ1), are localized on the outer mitochondrial membrane, referred to as mitoAMPK, in various tissues in mice and humans. Activation of mitoAMPK varies across the reticulum in response to energetic stress, and inhibition of mitoAMPK activity attenuates exercise-induced mitophagy in skeletal muscle in vivo. Discovery of a mitochondrial pool of AMPK and its local importance for mitochondrial quality control underscores the complexity of sensing cellular energetics in vivo that has implications for targeting mitochondrial energetics for disease treatment.
    Keywords:  AMPK; exercise; mitochondria; mitophagy; skeletal muscle
  21. Proc Natl Acad Sci U S A. 2021 Sep 14. pii: e2011226118. [Epub ahead of print]118(37):
      Alzheimer's disease (AD) is a progressive and fatal neurodegenerative disorder. Impaired neuronal bioenergetics and neuroinflammation are thought to play key roles in the progression of AD, but their interplay is not clear. Nicotinamide adenine dinucleotide (NAD+) is an important metabolite in all human cells in which it is pivotal for multiple processes including DNA repair and mitophagy, both of which are impaired in AD neurons. Here, we report that levels of NAD+ are reduced and markers of inflammation increased in the brains of APP/PS1 mutant transgenic mice with beta-amyloid pathology. Treatment of APP/PS1 mutant mice with the NAD+ precursor nicotinamide riboside (NR) for 5 mo increased brain NAD+ levels, reduced expression of proinflammatory cytokines, and decreased activation of microglia and astrocytes. NR treatment also reduced NLRP3 inflammasome expression, DNA damage, apoptosis, and cellular senescence in the AD mouse brains. Activation of cyclic GMP-AMP synthase (cGAS) and stimulator of interferon genes (STING) are associated with DNA damage and senescence. cGAS-STING elevation was observed in the AD mice and normalized by NR treatment. Cell culture experiments using microglia suggested that the beneficial effects of NR are, in part, through a cGAS-STING-dependent pathway. Levels of ectopic (cytoplasmic) DNA were increased in APP/PS1 mutant mice and human AD fibroblasts and down-regulated by NR. NR treatment induced mitophagy and improved cognitive and synaptic functions in APP/PS1 mutant mice. Our findings suggest a role for NAD+ depletion-mediated activation of cGAS-STING in neuroinflammation and cellular senescence in AD.
    Keywords:  Alzheimer’s disease; DNA repair; NAD supplementation; inflammation; neurodegeneration
  22. Front Mol Biosci. 2021 ;8 697359
      Declines in cellular nicotinamide adenine dinucleotide (NAD) contribute to metabolic dysfunction, increase susceptibility to disease, and occur as a result of pathogenic infection. The enzymatic cleavage of NAD+ transfers ADP-ribose (ADPr) to substrate proteins generating mono-ADP-ribose (MAR), poly-ADP-ribose (PAR) or O-acetyl-ADP-ribose (OAADPr). These important post-translational modifications have roles in both immune response activation and the advancement of infection. In particular, emergent data show viral infection stimulates activation of poly (ADP-ribose) polymerase (PARP) mediated NAD+ depletion and stimulates hydrolysis of existing ADP-ribosylation modifications. These studies are important for us to better understand the value of NAD+ maintenance upon the biology of infection. This review focuses specifically upon the NAD+ utilising enzymes, discusses existing knowledge surrounding their roles in infection, their NAD+ depletion capability and their influence within pathogenic infection.
    Keywords:  ARTCs; Infectious disease; NAD+; PARPs; SARM-1; SIRTs; cADPr
  23. J Proteome Res. 2021 Sep 09.
      Human pluripotent stem-cell-derived cardiomyocytes (hPSC-CMs) show immense promise for patient-specific disease modeling, cardiotoxicity screening, and regenerative therapy development. However, thus far, hPSC-CMs in culture have not recapitulated the structural or functional properties of adult CMs in vivo. To gain global insight into hPSC-CM biology, we established a multiomics method for analyzing the hPSC-CM metabolome and proteome from the same cell culture, creating multidimensional profiles of hPSC-CMs. Specifically, we developed a sequential extraction to capture metabolites and proteins from the same hPSC-CM monolayer cultures and analyzed these extracts using high-resolution mass spectrometry. Using this method, we annotated 205 metabolites/lipids and 4319 proteins from 106 cells with high reproducibility. We further integrated the proteome and metabolome measurements to create network profiles of molecular phenotypes for hPSC-CMs. Out of 310 pathways identified using metabolomics and proteomics, 40 pathways were considered significantly overrepresented (false-discovery-rate-corrected p ≤ 0.05). Highly populated pathways included those involved in protein synthesis (ribosome, spliceosome), ATP generation (oxidative phosphorylation), and cardiac muscle contraction. This multiomics method achieves a deep coverage of metabolites and proteins, creating a multidimensional view of the hPSC-CM phenotype, which provides a strong technological foundation to advance the understanding of hPSC-CM biology. Raw data are available in the MassIVE repository with identifier MSV000088010.
    Keywords:  FTICR MS; Multiomics; bottom-up proteomics; cell culture extraction; stem-cell-derived cardiomyocytes; timsTOF Pro; untargeted metabolomics
  24. Brief Bioinform. 2021 Sep 08. pii: bbab363. [Epub ahead of print]
      Rare diseases occur in a smaller proportion of the general population, which is variedly defined as less than 200 000 individuals (US) or in less than 1 in 2000 individuals (Europe). Although rare, they collectively make up to approximately 7000 different disorders, with majority having a genetic origin, and affect roughly 300 million people globally. Most of the patients and their families undergo a long and frustrating diagnostic odyssey. However, advances in the field of genomics have started to facilitate the process of diagnosis, though it is hindered by the difficulty in genome data analysis and interpretation. A major impediment in diagnosis is in the understanding of the diverse approaches, tools and datasets available for variant prioritization, the most important step in the analysis of millions of variants to select a few potential variants. Here we present a review of the latest methodological developments and spectrum of tools available for rare disease genetic variant discovery and recommend appropriate data interpretation methods for variant prioritization. We have categorized the resources based on various steps of the variant interpretation workflow, starting from data processing, variant calling, annotation, filtration and finally prioritization, with a special emphasis on the last two steps. The methods discussed here pertain to elucidating the genetic basis of disease in individual patient cases via trio- or family-based analysis of the genome data. We advocate the use of a combination of tools and datasets and to follow multiple iterative approaches to elucidate the potential causative variant.
    Keywords:  bioinformatics tools; next-generation sequencing; rare diseases; trio-analysis; variant analysis; variant annotation; variant filtration; variant interpretation; variant prioritization; whole genome sequencing
  25. Front Genet. 2021 ;12 698825
      Background: The triad of drug efficacy, toxicity and resistance underpins the risk-benefit balance of all therapeutics. The application of pharmacogenomics has the potential to improve the risk-benefit balance of a given therapeutic via the stratification of patient populations based on DNA variants. A growth in the understanding of the particulars of the mitochondrial genome, alongside the availability of techniques for its interrogation has resulted in a growing body of literature examining the impact of mitochondrial DNA (mtDNA) variation upon drug response. Objective: To critically evaluate and summarize the available literature, across a defined period, in a systematic fashion in order to map out the current landscape of the subject area and identify how the field may continue to advance. Methods: A systematic review of the literature published between January 2009 and December 2020 was conducted using the PubMed database with the following key inclusion criteria: reference to specific mtDNA polymorphisms or haplogroups, a core objective to examine associations between mtDNA variants and drug response, and research performed using human subjects or human in vitro models. Results: Review of the literature identified 24 articles reporting an investigation of the association between mtDNA variant(s) and drug efficacy, toxicity or resistance that met the key inclusion criteria. This included 10 articles examining mtDNA variations associated with antiretroviral therapy response, 4 articles examining mtDNA variants associated with anticancer agent response and 4 articles examining mtDNA variants associated with antimicrobial agent response. The remaining articles covered a wide breadth of medications and were therefore grouped together and referred to as "other." Conclusions: Investigation of the impact of mtDNA variation upon drug response has been sporadic to-date. Collective assessment of the associations identified in the articles was inconclusive due to heterogeneous methods and outcomes, limited racial/ethnic groups, lack of replication and inadequate statistical power. There remains a high degree of idiosyncrasy in drug response and this area has the potential to explain variation in drug response in a clinical setting, therefore further research is likely to be of clinical benefit.
    Keywords:  drug; efficacy; haplogroup; mitochondrial DNA; resistance; response; toxicity
  26. Cell Rep. 2021 Sep 07. pii: S2211-1247(21)01112-8. [Epub ahead of print]36(10): 109668
      Aging, genomic stress, and mitochondrial dysfunction are risk factors for neurodegenerative pathologies, such as Parkinson disease (PD). Although genomic instability is associated with aging and mitochondrial impairment, the underlying mechanisms are poorly understood. Here, we show that base excision repair generates genomic stress, promoting age-related neurodegeneration in a Caenorhabditis elegans PD model. A physiological level of NTH-1 DNA glycosylase mediates mitochondrial and nuclear genomic instability, which promote degeneration of dopaminergic neurons in older nematodes. Conversely, NTH-1 deficiency protects against α-synuclein-induced neurotoxicity, maintaining neuronal function with age. This apparent paradox is caused by modulation of mitochondrial transcription in NTH-1-deficient cells, and this modulation activates LMD-3, JNK-1, and SKN-1 and induces mitohormesis. The dependance of neuroprotection on mitochondrial transcription highlights the integration of BER and transcription regulation during physiological aging. Finally, whole-exome sequencing of genomic DNA from patients with idiopathic PD suggests that base excision repair might modulate susceptibility to PD in humans.
    Keywords:  C. elegans; DNA-glycosylase; NTH-1; Parkinson disease; aging; base excision repair; hydrogen peroxide; mitohormesis; neurodegeneration; oxidative DNA damage
  27. Mol Psychiatry. 2021 Sep 06.
      Respiratory chain complex I deficiency elicits mitochondrial dysfunction and reactive oxidative species (ROS), which plays a crucial role in Parkinson's disease (PD) pathogenesis. However, it remains unclear whether the impairment in other complexes in the mitochondrial oxidative phosphorylation chain is also sufficient to trigger PD onset. Here we show that inhibition of Complex II or III in the electron transport chain (ETC) induces the motor disorder and PD pathologies in neuronal Thy1-C/EBPβ transgenic mice. Through a cell-based screening of mitochondrial respiratory chain inhibitors, we identified TTFA (complex II inhibitor) and Atovaquone (complex III inhibitor), which robustly block the oxidative phosphorylation functions, strongly escalate ROS, and activate C/EBPβ/AEP pathway that triggers dopaminergic neuronal cell death. Oral administration of these inhibitors to Thy1-C/EBPβ mice elicits constipation and motor defects, associated with Lewy body-like inclusions. Deletion of SDHD (Succinate dehydrogenase) gene from the complex II in the Substantia Nigra of Thy1-C/EBPβ mice triggers ROS and PD pathologies, resulting in motor disorders. Hence, our findings demonstrate that mitochondrial ETC inactivation triggers PD pathogenesis via activating C/EBPβ/AEP pathway.
  28. Mol Ther Nucleic Acids. 2021 Sep 03. 25 416-443
      Pathological cardiac hypertrophy begins as an adaptive response to increased workload; however, sustained hemodynamic stress will lead it to maladaptation and eventually cardiac failure. Mitochondria, being the powerhouse of the cells, can regulate cardiac hypertrophy in both adaptive and maladaptive phases; they are dynamic organelles that can adjust their number, size, and shape through a process called mitochondrial dynamics. Recently, several studies indicate that promoting mitochondrial fusion along with preventing mitochondrial fission could improve cardiac function during cardiac hypertrophy and avert its progression toward heart failure. However, some studies also indicate that either hyperfusion or hypo-fission could induce apoptosis and cardiac dysfunction. In this review, we summarize the recent knowledge regarding the effects of mitochondrial dynamics on the development and progression of cardiac hypertrophy with particular emphasis on the regulatory role of mitochondrial dynamics proteins through the genetic, epigenetic, and post-translational mechanisms, followed by discussing the novel therapeutic strategies targeting mitochondrial dynamic pathways.
    Keywords:  cardiac hypertrophy; mitochondrial dynamics; molecular mechanism; recent advances; therapeutic potential
  29. J Biol Chem. 2021 Sep 06. pii: S0021-9258(21)00976-5. [Epub ahead of print] 101174
      Mitochondrial Ca2+ uptake tailors the strength of stimulation of plasma membrane phospholipase C-coupled receptors to that of cellular bioenergetics. However, how Ca2+ uptake by the mitochondrial Ca2+ uniporter (MCU) shapes receptor-evoked inter-organellar Ca2+ signaling is unknown. Here, we used CRISPR/Cas9 gene knockout, sub-cellular Ca2+ imaging and mathematical modeling to show that MCU is a universal regulator of intracellular Ca2+ signaling across mammalian cell types. MCU activity sustains cytosolic Ca2+ signaling by preventing Ca2+-dependent inactivation (CDI) of store-operated Ca2+ release-activated Ca2+ (CRAC) channels and by inhibiting Ca2+ extrusion. Paradoxically, MCU knockout (MCU-KO) enhanced cytosolic Ca2+ responses to store depletion. Physiological agonist stimulation in MCU-KO cells led to enhanced frequency of cytosolic Ca2+ oscillations, endoplasmic reticulum (ER) Ca2+ refilling, nuclear translocation of nuclear factor for activated T-cells (NFAT) transcription factors, and cell proliferation, without altering inositol-1,4,5-trisphosphate receptor activity. Our data show that MCU has dual counterbalancing functions at the cytosol-mitochondria interface, whereby the cell-specific MCU-dependent cytosolic Ca2+ clearance and buffering capacity of mitochondria reciprocally regulate inter-organellar Ca2+ transfer and NFAT nuclear translocation during receptor-evoked signaling. These findings highlight the critical dual function of the MCU not only in the acute Ca2+ buffering by mitochondria but also in shaping ER and cytosolic Ca2+ signals that regulate cellular transcription and function.
    Keywords:  CRAC channels; MCU; NFAT; SOCE; calcium oscillations; calcium signaling; mitochondria
  30. Cell Metab. 2021 Sep 07. pii: S1550-4131(21)00371-5. [Epub ahead of print]33(9): 1723-1725
      Mitochondria have emerged as critical players in immune homeostasis and disease. Recently in Cell, while characterizing the removal of mitochondria from red blood cells during human erythropoiesis, Caielli et al. highlight the interferon-inducing potential of erythrocyte-derived mitochondria in lupus.
  31. Anal Chem. 2021 Sep 10.
      Mitochondrial pH (pHmito) is intimately related to mitochondrial function, and aberrant values for pHmito are linked to several disease states. We report the design, synthesis, and application of mitokyne 1-the first small molecule pHmito sensor for stimulated Raman scattering (SRS) microscopy. This ratiometric probe can determine subtle changes in pHmito in response to external stimuli and the inhibition of both the electron transport chain and ATP synthase with small molecule inhibitors. In addition, 1 was also used to monitor mitochondrial dynamics in a time-resolved manner with subcellular spatial resolution during mitophagy providing a powerful tool for dissecting the molecular and cell biology of this critical organelle.
  32. Nat Commun. 2021 09 06. 12(1): 5276
      A promise of genomics in precision medicine is to provide individualized genetic risk predictions. Polygenic risk scores (PRS), computed by aggregating effects from many genomic variants, have been developed as a useful tool in complex disease research. However, the application of PRS as a tool for predicting an individual's disease susceptibility in a clinical setting is challenging because PRS typically provide a relative measure of risk evaluated at the level of a group of people but not at individual level. Here, we introduce a machine-learning technique, Mondrian Cross-Conformal Prediction (MCCP), to estimate the confidence bounds of PRS-to-disease-risk prediction. MCCP can report disease status conditional probability value for each individual and give a prediction at a desired error level. Moreover, with a user-defined prediction error rate, MCCP can estimate the proportion of sample (coverage) with a correct prediction.
  33. Elife. 2021 Sep 06. pii: e69310. [Epub ahead of print]10
      Background: Bicuspid aortic valve (BAV) is the most common congenital cardiovascular disease in general population and is frequently associated with the development of thoracic aortic aneurysm (TAA). There is no effective strategy to intervene with TAA progression due to an incomplete understanding of the pathogenesis. Insufficiency of NOTCH1 expression is highly related to BAV-TAA, but the underlying mechanism remains to be clarified.
    Methods: A comparative proteomics analysis was used to explore the biological differences between non-diseased and BAV-TAA aortic tissues. A microfluidics-based aorta smooth muscle-on-a-chip model was constructed to evaluate the effect of NOTCH1 deficiency on contractile phenotype and mitochondrial dynamics of human aortic smooth muscle cells (HAoSMCs).
    Results: Protein analyses of human aortic tissues showed the insufficient expression of NOTCH1 and impaired mitochondrial dynamics in BAV-TAA. HAoSMCs with NOTCH1-knockdown exhibited reduced contractile phenotype and were accompanied by attenuated mitochondrial fusion. Furthermore, we identified that mitochondrial fusion activators (leflunomide and teriflunomide) or mitochondrial fission inhibitor (Mdivi-1) partially rescued the disorders of mitochondrial dynamics in HAoSMCs derived from BAV-TAA patients.
    Conclusions: The aorta smooth muscle-on-a-chip model simulates the human pathophysiological parameters of aorta biomechanics and provides a platform for molecular mechanism studies of aortic disease and related drug screening. This aorta smooth muscle-on-a-chip model and human tissue proteomic analysis revealed that impaired mitochondrial dynamics could be a potential therapeutic target for BAV-TAA.
    Funding: National Key R&D Program of China, National Natural Science Foundation of China, Shanghai Municipal Science and Technology Major Project, Shanghai Science and Technology Commission, and Shanghai Municipal Education Commission.
    Keywords:  human; medicine
  34. Nat Commun. 2021 09 06. 12(1): 5277
      The pyruvate dehydrogenase complex (PDHc) links glycolysis to the citric acid cycle by converting pyruvate into acetyl-coenzyme A. PDHc encompasses three enzymatically active subunits, namely pyruvate dehydrogenase, dihydrolipoyl transacetylase, and dihydrolipoyl dehydrogenase. Dihydrolipoyl transacetylase is a multidomain protein comprising a varying number of lipoyl domains, a peripheral subunit-binding domain, and a catalytic domain. It forms the structural core of the complex, provides binding sites for the other enzymes, and shuffles reaction intermediates between the active sites through covalently bound lipoyl domains. The molecular mechanism by which this shuttling occurs has remained elusive. Here, we report a cryo-EM reconstruction of the native E. coli dihydrolipoyl transacetylase core in a resting state. This structure provides molecular details of the assembly of the core and reveals how the lipoyl domains interact with the core at the active site.
  35. Hum Mol Genet. 2021 Sep 09. pii: ddab265. [Epub ahead of print]
      Recently, we and others identified de novo FBXO11 variants as causative for a variable neurodevelopmental disorder (NDD). We now assembled clinical and mutational information on 23 additional individuals. The phenotypic spectrum remains highly variable, with developmental delay and/or intellectual disability as the core feature and behavioral anomalies, hypotonia and various facial dysmorphism as frequent aspects. The mutational spectrum includes intragenic deletions, likely gene disrupting and missense variants distributed across the protein. To further characterize the functional consequences of FBXO11 missense variants, we analyzed their effects on protein expression and localization by overexpression of 17 different mutant constructs in HEK293 and HeLa cells. We found that the majority of missense variants resulted in subcellular mislocalization and/or reduced FBXO11 protein expression levels. For instance, variants located in the nuclear localization signal and the N-terminal F-Box domain lead to altered subcellular localization with exclusion from the nucleus or the formation of cytoplasmic aggregates and to reduced protein levels in western blot. In contrast, variants localized in the C-terminal Zn-finger UBR domain lead to an accumulation in the cytoplasm without alteration of protein levels. Together with the mutational data our functional results suggest that most missense variants likely lead to a loss of the original FBXO11 function and thereby highlight haploinsufficiency as the most likely disease mechanism for FBXO11-associated NDDs.
  36. Eur J Hum Genet. 2021 Sep 06.
      Mitochondrial disorders are a heterogeneous group of rare, degenerative multisystem disorders affecting the cell's core bioenergetic and signalling functions. Spontaneous improvement is rare. We describe a novel neonatal-onset mitochondriopathy in three infants with failure to thrive, hyperlactatemia, hyperammonemia, and apparent clinical resolution before 18 months. Exome sequencing showed all three probands to be identically heterozygous for a recurrent de novo substitution, c.620G>A [p.(Arg207His)] in ATP5F1A, encoding the α-subunit of complex V. Patient-derived fibroblasts exhibited multiple deficits in complex V function and expression in vitro. Structural modelling predicts the observed substitution to create an abnormal region of negative charge on ATP5F1A's β-subunit-interacting surface, adjacent to the nearby β subunit's active site. This disorder, which presents with life-threatening neonatal manifestations, appears to follow a remitting course; the long-term prognosis remains unknown.
  37. Eur J Pharm Biopharm. 2021 Sep 06. pii: S0939-6411(21)00233-2. [Epub ahead of print]
      Leber's Hereditary Optic Neuropathy (LHON) is a hereditary mitochondrial neurodegenerative disease of unclear etiology and lack of available therapeutic alternatives. The main goal of the current pilot study was based on the evaluation of the feasibility and characteristics of prolonged and controlled idebenone release from a PCL intravitreal implant. The design, development, and characterization of idebenone-loaded PCL implants prepared by an homogenization/extrusion/solvent evaporation method allowed the obtention of high PY, EE and LC values. In vitro characterization was completed by the assessment of mechanical and instrumental properties. The in vitro release of idebenone from the PCL implants was assessed and the implant erosion was monitored by the mass loss and surface morphology changes. DSC was used to estimate stability and interaction among implant's components. The present work demonstrated the controlled and prolonged idebenone delivery from the PCL implants in an in vitro model. A consistent preclinical base was established, supporting the idea of idebenone-loaded PCL implants as a new strategy of long-term sustained intraocular delivery for the LHON treatment.
    Keywords:  LHON; controlled drug delivery; idebenone; intravitreal administration; poly-ε-caprolactone; polymeric implant