Signal transducer and activator of transcription 2 deficiency is a novel disorder of mitochondrial fission

Defects of mitochondrial dynamics are emerging causes of neurological disease. In two children presenting with severe neurological deterioration following viral infection we identified a novel homozygous STAT2 mutation, c.1836C4A (p.Cys612Ter), using whole exome sequencing. In muscle and fibroblasts from these patients, and a third unrelated STAT2-deficient patient, we observed extremely elongated mitochondria. Western blot analysis revealed absence of the STAT2 protein and that the mitochondrial fission protein DRP1 (encoded by DNM1L) is inactive, as shown by its phosphorylation state. All three patients harboured 15 decreased levels of DRP1 phosphorylated at serine residue 616 (P-DRP1S616), a post-translational modification known to activate DRP1, and increased levels of DRP1 phosphorylated at serine 637 (P-DRP1S637), associated with the inactive state of the DRP1 GTPase. Knockdown of STAT2 in SHSY5Y cells recapitulated the fission defect, with elongated mitochondria and decreased PDRP1 S616 levels. Furthermore the mitochondrial fission defect in patient fibroblasts was rescued following lentiviral transduction with wild-type STAT2 in all three patients, with normalization of mitochondrial length and increased P-DRP1S616 levels. Taken 20 together, these findings implicate STAT2 as a novel regulator of DRP1 phosphorylation at serine 616, and thus of mitochondrial fission, and suggest that there are interactions between immunity and mitochondria. This is the first study to link the innate immune system to mitochondrial dynamics and morphology. We hypothesize that variability in JAK-STAT signalling may contribute to the phenotypic heterogeneity of mitochondrial disease, and may explain why some patients with underlying mitochondrial disease decompensate after seemingly trivial viral infections. Modulating JAK-STAT activity may represent a novel 25 therapeutic avenue for mitochondrial diseases, which remain largely untreatable. This may also be relevant for more common neurodegenerative diseases, including Alzheimer’s, Huntington’s and Parkinson’s diseases, in which abnormalities of mitochondrial morphology have been implicated in disease pathogenesis

Loss of α-Synuclein Does Not Affect Mitochondrial Bioenergetics in Rodent Neurons.

Increased α-synuclein (αsyn) and mitochondrial dysfunction play central roles in the pathogenesis of Parkinson's disease (PD), and lowering αsyn is under intensive investigation as a therapeutic strategy for PD. Increased αsyn levels disrupt mitochondria and impair respiration, while reduced αsyn protects against mitochondrial toxins, suggesting that interactions between αsyn and mitochondria influences the pathologic and physiologic functions of αsyn. However, we do not know if αsyn affects normal mitochondrial function or if lowering αsyn levels impacts bioenergetic function, especially at the nerve terminal where αsyn is enriched. To determine if αsyn is required for normal mitochondrial function in neurons, we comprehensively evaluated how lowering αsyn affects mitochondrial function. We found that αsyn knockout (KO) does not affect the respiration of cultured hippocampal neurons or cortical and dopaminergic synaptosomes, and that neither loss of αsyn nor all three (α, β and γ) syn isoforms decreased mitochondria-derived ATP levels at the synapse. Similarly, neither αsyn KO nor knockdown altered the capacity of synaptic mitochondria to meet the energy requirements of synaptic vesicle cycling or influenced the localization of mitochondria to dopamine (DA) synapses in vivo. Finally, αsyn KO did not affect overall energy metabolism in mice assessed with a Comprehensive Lab Animal Monitoring System. These studies suggest either that αsyn has little or no significant physiological effect on mitochondrial bioenergetic function, or that any such functions are fully compensated for when lost. These results implicate that αsyn levels can be reduced in neurons without impairing (or improving) mitochondrial bioenergetics or distribution

TOM40 Mediates Mitochondrial Dysfunction Induced by α-Synuclein Accumulation in Parkinson's Disease.

Alpha-synuclein (α-Syn) accumulation/aggregation and mitochondrial dysfunction play prominent roles in the pathology of Parkinson's disease. We have previously shown that postmortem human dopaminergic neurons from PD brains accumulate high levels of mitochondrial DNA (mtDNA) deletions. We now addressed the question, whether alterations in a component of the mitochondrial import machinery -TOM40- might contribute to the mitochondrial dysfunction and damage in PD. For this purpose, we studied levels of TOM40, mtDNA deletions, oxidative damage, energy production, and complexes of the respiratory chain in brain homogenates as well as in single neurons, using laser-capture-microdissection in transgenic mice overexpressing human wildtype α-Syn. Additionally, we used lentivirus-mediated stereotactic delivery of a component of this import machinery into mouse brain as a novel therapeutic strategy. We report here that TOM40 is significantly reduced in the brain of PD patients and in α-Syn transgenic mice. TOM40 deficits were associated with increased mtDNA deletions and oxidative DNA damage, and with decreased energy production and altered levels of complex I proteins in α-Syn transgenic mice. Lentiviral-mediated overexpression of Tom40 in α-Syn-transgenic mice brains ameliorated energy deficits as well as oxidative burden. Our results suggest that alterations in the mitochondrial protein transport machinery might contribute to mitochondrial impairment in α-Synucleinopathies

Highly efficient 5\u27 capping of mitochondrial RNA with NAD+ and NADH by yeast and human mitochondrial RNA polymerase

Bacterial and eukaryotic nuclear RNA polymerases (RNAPs) cap RNA with the oxidized and reduced forms of the metabolic effector nicotinamide adenine dinucleotide, NAD+ and NADH, using NAD+ and NADH as non-canonical initiating nucleotides for transcription initiation. Here, we show that mitochondrial RNAPs (mtRNAPs) cap RNA with NAD+ and NADH, and do so more efficiently than nuclear RNAPs. Direct quantitation of NAD+- and NADH-capped RNA demonstrates remarkably high levels of capping in vivo: up to ~60% NAD+ and NADH capping of yeast mitochondrial transcripts, and up to ~15% NAD+ capping of human mitochondrial transcripts. The capping efficiency is determined by promoter sequence at, and upstream of, the transcription start site and, in yeast and human cells, by intracellular NAD+ and NADH levels. Our findings indicate mtRNAPs serve as both sensors and actuators in coupling cellular metabolism to mitochondrial transcriptional outputs, sensing NAD+ and NADH levels and adjusting transcriptional outputs accordingly. © 2018, Bird et al

Mitochondrial dysfunction promoted by Porphyromonas gingivalis lipopolysaccharide as a possible link between cardiovascular disease and periodontitis

Oxidative stress is one of the factors that could explain the pathophysiological mechanism of inflammatory conditions that occur in cardiovascular disease (CVD) and periodontitis. Such inflammatory response is often evoked by specific bacteria, as the lipopolysaccharide (LPS) of Porphyromonas gingivalis is a key factor in this process. The aim of this research was to study the role of mitochondrial dysfunction in peripheral blood mononuclear cells (PBMCs) from periodontitis patients and to evaluate the influence of LPS on fibroblasts to better understand the pathophysiology of periodontitis and its relationship with CVD. PBMCs from patients showed lower CoQ10 levels and citrate synthase activity, together with high levels of ROS production. LPS-treated fibroblasts provoked increased oxidative stress and mitochondrial dysfunction by a decrease in mitochondrial protein expression, mitochondrial mass, and mitochondrial membrane potential. Our study supports the hypothesis that LPS-mediated mitochondrial dysfunction could be at the origin of oxidative stress in periodontal patients. Abnormal PBMC performance may promote oxidative stress and alter cytokine homeostasis. In conclusion, mitochondrial dysfunction could represent a possible link to understanding the interrelationships between two prominent inflammatory diseases: periodontitis and CV

Inhibition of the mitochondrial pyruvate carrier protects from excitotoxic neuronal death.

Glutamate is the dominant excitatory neurotransmitter in the brain, but under conditions of metabolic stress it can accumulate to excitotoxic levels. Although pharmacologic modulation of excitatory amino acid receptors is well studied, minimal consideration has been given to targeting mitochondrial glutamate metabolism to control neurotransmitter levels. Here we demonstrate that chemical inhibition of the mitochondrial pyruvate carrier (MPC) protects primary cortical neurons from excitotoxic death. Reductions in mitochondrial pyruvate uptake do not compromise cellular energy metabolism, suggesting neuronal metabolic flexibility. Rather, MPC inhibition rewires mitochondrial substrate metabolism to preferentially increase reliance on glutamate to fuel energetics and anaplerosis. Mobilizing the neuronal glutamate pool for oxidation decreases the quantity of glutamate released upon depolarization and, in turn, limits the positive-feedback cascade of excitotoxic neuronal injury. The finding links mitochondrial pyruvate metabolism to glutamatergic neurotransmission and establishes the MPC as a therapeutic target to treat neurodegenerative diseases characterized by excitotoxicity

Evidence for variation in the effective population size of animal mitochondrial DNA

Background: It has recently been shown that levels of diversity in mitochondrial DNA are remarkably constant across animals of diverse census population sizes and ecologies, which has led to the suggestion that the effective population of mitochondrial DNA may be relatively constant. Results: Here we present several lines of evidence that suggest, to the contrary, that the effective population size of mtDNA does vary, and that the variation can be substantial. First, we show that levels of mitochondrial and nuclear diversity are correlated within all groups of animals we surveyed. Second, we show that the effectiveness of selection on non-synonymous mutations, as measured by the ratio of the numbers of non-synonymous and synonymous polymorphisms, is negatively correlated to levels of mitochondrial diversity. Finally, we estimate the effective population size of mitochondrial DNA in selected mammalian groups and show that it varies by at least an order of magnitude. Conclusions: We conclude that there is variation in the effective population size of mitochondria. Furthermore we suggest that the relative constancy of DNA diversity may be due to a negative correlation between the effective population size and the mutation rate per generation

Mutations in valosin-containing protein (VCP) decrease ADP/ATP translocation across the mitochondrial membrane and impair energy metabolism in human neurons

Mutations in the gene encoding valosin-containing protein (VCP) lead to multisystem proteinopathies including frontotemporal dementia. We have previously shown that patient-derived VCP mutant fibroblasts exhibit lower mitochondrial membrane potential, uncoupled respiration, and reduced ATP levels. This study addresses the underlying basis for mitochondrial uncoupling using VCP knockdown neuroblastoma cell lines, induced pluripotent stem cells (iPSCs), and iPSC-derived cortical neurons from patients with pathogenic mutations in VCP. Using fluorescent live cell imaging and respiration analysis we demonstrate a VCP mutation/knockdown-induced dysregulation in the adenine nucleotide translocase, which results in a slower rate of ADP or ATP translocation across the mitochondrial membranes. This deregulation can explain the mitochondrial uncoupling and lower ATP levels in VCP mutation-bearing neurons via reduced ADP availability for ATP synthesis. This study provides evidence for a role of adenine nucleotide translocase in the mechanism underlying altered mitochondrial function in VCP-related degeneration, and this new insight may inform efforts to better understand and manage neurodegenerative disease and other proteinopathies

Mutant mitochondrial elongation factor G1 and combined oxidative phosphorylation deficiency

Although most components of the mitochondrial translation apparatus are encoded by nuclear genes, all known molecular defects associated with impaired mitochondrial translation are due to mutations in mitochondrial DNA. We investigated two siblings with a severe defect in mitochondrial translation, reduced levels of oxidative phosphorylation complexes containing mitochondrial DNA (mtDNA)–encoded subunits, and progressive hepatoencephalopathy. We mapped the defective gene to a region on chromosome 3q containing elongation factor G1 (EFG1), which encodes a mitochondrial translation factor. Sequencing of EFG1 revealed a mutation affecting a conserved residue of the guanosine triphosphate (GTP)–binding domain. These results define a new class of gene defects underlying disorders of oxidative phosphorylation