Littérature scientifique sur le sujet « Ataxia, SCA28, Neurodegeneration, mitochondria »

BackgroundSpinocerebellar ataxia type 28 (SCA28) is a dominantly inherited neurodegenerative disease caused by pathogenic variants in AFG3L2. The AFG3L2 protein is a subunit of mitochondrial m-AAA complexes involved in protein quality control. Objective of this study was to determine the molecular mechanisms of SCA28, which has eluded characterisation to date.MethodsWe derived SCA28 patient fibroblasts carrying different pathogenic variants in the AFG3L2 proteolytic domain (missense: the newly identified p.F664S and p.M666T, p.G671R, p.Y689H and a truncating frameshift p.L556fs) and analysed multiple aspects of mitochondrial physiology. As reference of residual m-AAA activity, we included SPAX5 patient fibroblasts with homozygous p.Y616C pathogenic variant, AFG3L2+/− HEK293 T cells by CRISPR/Cas9-genome editing and Afg3l2−/− murine fibroblasts.ResultsWe found that SCA28 cells carrying missense changes have normal levels of assembled m-AAA complexes, while the cells with a truncating pathogenic variant had only half of this amount. We disclosed inefficient mitochondrial fusion in SCA28 cells caused by increased OPA1 processing operated by hyperactivated OMA1. Notably, we found altered mitochondrial proteostasis to be the trigger of OMA1 activation in SCA28 cells, with pharmacological attenuation of mitochondrial protein synthesis resulting in stabilised levels of OMA1 and OPA1 long forms, which rescued mitochondrial fusion efficiency. Secondary to altered mitochondrial morphology, mitochondrial calcium uptake resulted decreased in SCA28 cells.ConclusionOur data identify the earliest events in SCA28 pathogenesis and open new perspectives for therapy. By identifying similar mitochondrial phenotypes between SCA28 cells and AFG3L2+/− cells, our results support haploinsufficiency as the mechanism for the studied pathogenic variants.

A role for mitochondrial dysfunction in neurodegenerative disease is gaining increasing support. Mitochondrial dysfunction may be linked to neurodegenerative diseases through a variety of different pathways, including free-radical generation, impaired calcium buffering and the mitochondrial permeability transition. This can lead to both apoptotic and necrotic cell death. Recent evidence has shown that there is a mitochondrial defect in Friedreich's ataxia, which leads to increased mitochondrial iron content, that appears to be linked to increased free-radical generation. There is evidence that the point mutations in superoxide dismutase which are associated with amyotrophic lateral sclerosis may contribute to mitochondrial dysfunction. There is also evidence for bioenergetic defects in Huntington's disease. Studies of cybrid cell lines have implicated mitochondrial defects in both Parkinson's disease and Alzheimer's disease. If mitochondrial dysfunction plays a role in neurodegenerative diseases then therapeutic strategies such as coenzyme Q10 and creatine may be useful in attempting to slow the disease process.

Friedreich’s ataxia is the most common inherited autosomal recessive ataxia and is characterized by progressive degeneration of the peripheral and central nervous systems and cardiomyopathy. This disease is caused by the silencing of theFXNgene and reduced levels of the encoded protein, frataxin. Frataxin is a mitochondrial protein that functions primarily in iron-sulfur cluster synthesis. This small protein with anα/βsandwich fold undergoes complex processing and imports into the mitochondria, generating isoforms with distinct N-terminal lengths which may underlie different functionalities, also in respect to oligomerization. Missense mutations in theFXNcoding region, which compromise protein folding, stability, and function, are found in 4% of FRDA heterozygous patients and are useful to understand how loss of functional frataxin impacts on FRDA physiopathology. In cells, frataxin deficiency leads to pleiotropic phenotypes, including deregulation of iron homeostasis and increased oxidative stress. Increasing amount of data suggest that oxidative stress contributes to neurodegeneration in Friedreich’s ataxia.

Abstract Peroxiredoxin 3 (PRDX3) belongs to a superfamily of peroxidases that function as protective antioxidant enzymes. Among the six isoforms (PRDX1–PRDX6), PRDX3 is the only protein exclusively localized to the mitochondria, which are the main source of reactive oxygen species. Excessive levels of reactive oxygen species are harmful to cells, inducing mitochondrial dysfunction, DNA damage, lipid and protein oxidation and ultimately apoptosis. Neuronal cell damage induced by oxidative stress has been associated with numerous neurodegenerative disorders including Alzheimer’s and Parkinson’s diseases. Leveraging the large aggregation of genomic ataxia datasets from the PREPARE (Preparing for Therapies in Autosomal Recessive Ataxias) network, we identified recessive mutations in PRDX3 as the genetic cause of cerebellar ataxia in five unrelated families, providing further evidence for oxidative stress in the pathogenesis of neurodegeneration. The clinical presentation of individuals with PRDX3 mutations consists of mild-to-moderate progressive cerebellar ataxia with concomitant hyper- and hypokinetic movement disorders, severe early-onset cerebellar atrophy, and in part olivary and brainstem degeneration. Patient fibroblasts showed a lack of PRDX3 protein, resulting in decreased glutathione peroxidase activity and decreased mitochondrial maximal respiratory capacity. Moreover, PRDX3 knockdown in cerebellar medulloblastoma cells resulted in significantly decreased cell viability, increased H2O2 levels and increased susceptibility to apoptosis triggered by reactive oxygen species. Pan-neuronal and pan-glial in vivo models of Drosophila revealed aberrant locomotor phenotypes and reduced survival times upon exposure to oxidative stress. Our findings reveal a central role for mitochondria and the implication of oxidative stress in PRDX3 disease pathogenesis and cerebellar vulnerability and suggest targets for future therapeutic approaches.

Calcium (Ca2+) is a versatile secondary messenger involved in the regulation of a plethora of different signaling pathways for cell maintenance. Specifically, intracellular Ca2+ homeostasis is mainly regulated by the endoplasmic reticulum and the mitochondria, whose Ca2+ exchange is mediated by appositions, termed endoplasmic reticulum–mitochondria-associated membranes (MAMs), formed by proteins resident in both compartments. These tethers are essential to manage the mitochondrial Ca2+ influx that regulates the mitochondrial function of bioenergetics, mitochondrial dynamics, cell death, and oxidative stress. However, alterations of these pathways lead to the development of multiple human diseases, including neurological disorders, such as amyotrophic lateral sclerosis, Friedreich’s ataxia, and Charcot–Marie–Tooth. A common hallmark in these disorders is mitochondrial dysfunction, associated with abnormal mitochondrial Ca2+ handling that contributes to neurodegeneration. In this work, we highlight the importance of Ca2+ signaling in mitochondria and how the mechanism of communication in MAMs is pivotal for mitochondrial maintenance and cell homeostasis. Lately, we outstand potential targets located in MAMs by addressing different therapeutic strategies focused on restoring mitochondrial Ca2+ uptake as an emergent approach for neurological diseases.

DNA repair ensures genomic stability to achieve healthy ageing, including cognitive maintenance. Mutations on genes encoding key DNA repair proteins can lead to diseases with accelerated ageing phenotypes. Some of these diseases are xeroderma pigmentosum group A (XPA, caused by mutation of XPA), Cockayne syndrome group A and group B (CSA, CSB, and are caused by mutations of CSA and CSB, respectively), ataxia-telangiectasia (A-T, caused by mutation of ATM), and Werner syndrome (WS, with most cases caused by mutations in WRN). Except for WS, a common trait of the aforementioned progerias is neurodegeneration. Evidence from studies using animal models and patient tissues suggests that the associated DNA repair deficiencies lead to depletion of cellular nicotinamide adenine dinucleotide (NAD+), resulting in impaired mitophagy, accumulation of damaged mitochondria, metabolic derailment, energy deprivation, and finally leading to neuronal dysfunction and loss. Intriguingly, these features are also observed in Alzheimer’s disease (AD), the most common type of dementia affecting more than 50 million individuals worldwide. Further studies on the mechanisms of the DNA repair deficient premature ageing diseases will help to unveil the mystery of ageing and may provide novel therapeutic strategies for AD.

Abstract Ataxia-telangiectasia (AT) and MRE11-defective Ataxia-telangiectasia-like disorder (ATLD) patients show progressive cerebellar ataxia. ATM, mutated in AT, can be activated in response to oxidative stress as well as DNA damage, which could be linked to disease-related neurodegeneration. However, the role of MRE11 in oxidative stress responses has been elusive. Here, we showed that MRE11 could participate in ATM activation during oxidative stress in an NBS1/RAD50-independent manner. Importantly, MRE11 was indispensable for ATM activation. We identified FXR1 as a novel MRE11-binding partner by mass spectrometry. We confirmed that FXR1 could bind with MRE11 and showed that both localize to the cytoplasm. Notably, MRE11 and FXR1 partly localize to the mitochondria, which are the major source of cytoplasmic reactive oxygen species (ROS). The contribution of FXR1 to DNA double-strand break damage responses seemed minor and limited to HR repair, considering that depletion of FXR1 perturbed chromatin association of homologous recombination repair factors and sensitized cells to camptothecin. During oxidative stress, depletion of FXR1 by siRNA reduced oxidative stress responses and increased the sensitivity to pyocyanin, a mitochondrial ROS inducer. Collectively, our findings suggest that MRE11 and FXR1 might contribute to cellular defense against mitochondrial ROS as a cytoplasmic complex.

In 1988, the gene responsible for the autosomal recessive disease ataxia- telangiectasia (A-T) was localized to 11q22.3-23.1. It was eventually cloned in 1995. Many independent laboratories have since demonstrated that in replicating cells, ataxia telangiectasia mutated (ATM) is predominantly a nuclear protein that is involved in the early recognition and response to double-stranded DNA breaks. ATM is a high-molecular-weight PI3K-family kinase. ATM also plays many important cytoplasmic roles where it phosphorylates hundreds of protein substrates that activate and coordinate cell-signaling pathways involved in cell-cycle checkpoints, nuclear localization, gene transcription and expression, the response to oxidative stress, apoptosis, nonsense-mediated decay, and others. Appreciating these roles helps to provide new insights into the diverse clinical phenotypes exhibited by A-T patients—children and adults alike—which include neurodegeneration, high cancer risk, adverse reactions to radiation and chemotherapy, pulmonary failure, immunodeficiency, glucose transporter aberrations, insulin-resistant diabetogenic responses, and distinct chromosomal and chromatin changes. An exciting recent development is the ATM-dependent pathology encountered in mitochondria, leading to inefficient respiration and energy metabolism and the excessive generation of free radicals that themselves create life-threatening DNA lesions that must be repaired within minutes to minimize individual cell losses.

Mitophagy is a cellular process by which dysfunctional mitochondria are degraded via autophagy. Increasing empirical evidence proposes that this mitochondrial quality-control mechanism is defective in neurons of patients with various neurodegenerative diseases such as Ataxia Telangiectasia, Alzheimer’s disease, Parkinson’s disease, and Amyotrophic Lateral Sclerosis. Accumulation of defective mitochondria and the production of reactive oxygen species due to defective mitophagy have been identified as causes underlying neurodegenerative disease pathogenesis. However, the reason mitophagy is defective in most neurodegenerative diseases is unclear. Like mitophagy, defects in the ubiquitin/26S proteasome pathway have been linked to neurodegeneration, resulting in the characteristic protein aggregates often seen in neurons of affected patients. Although initiation of mitophagy requires a functional ubiquitin pathway, whether defects in the ubiquitin pathway are causally responsible for defective mitophagy is not known. In this mini-review, we introduce mitophagy and ubiquitin pathways and provide a summary of our current understanding of the regulation of mitophagy by the ubiquitin pathway. We will then briefly review empirical evidence supporting mitophagy defects in neurodegenerative diseases. The review will conclude with a discussion of the constitutively elevated expression of ubiquitin-like protein Interferon-Stimulated Gene 15 (ISG15), an antagonist of the ubiquitin pathway, as a potential cause of defective mitophagy in neurodegenerative diseases. Impact statement Neurodegenerative diseases place an enormous burden on patients and caregivers globally. Over six million people in the United States alone suffer from neurodegenerative diseases, all of which are chronic, incurable, and with causes unknown. Identifying a common molecular mechanism underpinning neurodegenerative disease pathology is urgently needed to aid in the design of effective therapies to ease suffering, reduce economic cost, and improve the quality of life for these patients. Although the development of neurodegeneration may vary between neurodegenerative diseases, they have common cellular hallmarks, including defects in the ubiquitin-proteasome system and mitophagy. In this review, we will provide a summary of our current understanding of the regulation of mitophagy by the ubiquitin pathway and discuss the potential of targeting mitophagy and ubiquitin pathways for the treatment of neurodegeneration.

Abstract Cerebral choline metabolism is crucial for normal brain function, and its homoeostasis depends on carrier-mediated transport. Here, we report on four individuals from three families with neurodegenerative disease and homozygous frameshift mutations (Asp517Metfs*19, Ser126Metfs*8, and Lys90Metfs*18) in the SLC44A1 gene encoding choline transporter-like protein 1. Clinical features included progressive ataxia, tremor, cognitive decline, dysphagia, optic atrophy, dysarthria, as well as urinary and bowel incontinence. Brain MRI demonstrated cerebellar atrophy and leukoencephalopathy. Moreover, low signal intensity in globus pallidus with hyperintensive streaking and low signal intensity in substantia nigra were seen in two individuals. The Asp517Metfs*19 and Ser126Metfs*8 fibroblasts were structurally and functionally indistinguishable. The most prominent ultrastructural changes of the mutant fibroblasts were reduced presence of free ribosomes, the appearance of elongated endoplasmic reticulum and strikingly increased number of mitochondria and small vesicles. When chronically treated with choline, those characteristics disappeared and mutant ultrastructure resembled healthy control cells. Functional analysis revealed diminished choline transport yet the membrane phosphatidylcholine content remained unchanged. As part of the mechanism to preserve choline and phosphatidylcholine, choline transporter deficiency was implicated in impaired membrane homeostasis of other phospholipids. Choline treatments could restore the membrane lipids, repair cellular organelles and protect mutant cells from acute iron overload. In conclusion, we describe a novel childhood-onset neurometabolic disease caused by choline transporter deficiency with autosomal recessive inheritance.

Autosomal dominant spinocerebellar ataxias (SCAs) are genetically heterogeneous neurological disorders characterized by cerebellar dysfunction mostly due to Purkinje cell degeneration. Here we show that AFG3L2 mutations cause SCA type 28. Along with paraplegin, which causes recessive spastic paraplegia, AFG3L2 is a component of the conserved m-AAA metalloprotease complex involved in the maintenance of the mitochondrial proteome. We identified heterozygous missense mutations in five unrelated SCA families and found that AFG3L2 is highly and selectively expressed in human cerebellar Purkinje cells. m-AAA–deficient yeast cells expressing human mutated AFG3L2 homocomplex show respiratory deficiency, proteolytic impairment and deficiency of respiratory chain complex IV. Structure homology modeling indicates that the mutations may affect AFG3L2 substrate handling. This work identifies AFG3L2 as a novel cause of dominant neurodegenerative disease and indicates a previously unknown role for this component of the mitochondrial protein quality control machinery in protecting the human cerebellum against neurodegeneration.

Autosomal dominant spinocerebellar ataxias (SCA) are a heterogeneous group of neurological disorders characterized by cerebellar dysfunction. We recently showed that AFG3L2 mutations cause dominant ataxia SCA28. AFG3L2 and its partner protein paraplegin, which causes recessive spastic paraparesis SPG7, are components of the m-AAA complex, involved in mitochondrial protein quality control. Since yeast functional studies showed that paraplegin coexpression can modulate AFG3L2 mutations, we investigated the possible coinheritance of AFG3L2 and SPG7 mutations in patients with spinocerebellar syndromes. We identified 3 probands with heterozygous mutations in both the AFG3L2 and the SPG7 genes. Two ataxic patients carry an AFG3L2 mutation affecting highly conserved amino acids located in the ATPase or in the proteolytic domains of the protein along with the parapleginA510V. The third proband carries a de novo AFG3L2 mutation in the highly conserved SRH region of the ATPase domain along with the inherited deletion of SPG7 exons 4-6. The clinical presentation of this patient is characterized by early onset optic atrophy and a L-dopa-responsive spastic-ataxic syndrome with extrapyramidal signs. A muscle biopsy revealed an isolated complex I deficiency. Moreover, evaluation of substrates processing in patient’s fibroblasts showed abnormal processing pattern of OPA1. In conclusion, our data indicate that the presence of a loss-of-function mutation in paraplegin may act as a disease modifier for heterozygous AFG3L2 mutations. Concurrent mutations in both components of the mitochondrial m-AAA complex may result in a complex phenotype, thus expanding the clinical spectrum of AFG3L2-associated mutations. Moreover, biochemical and cell biology studies revealed a crucial role of the m-AAA complex in the processing of OPA1 and the maintenance of mitochondrial morphology.

Friedreich’s ataxia (FA) is the most common autosomal recessive ataxia, and patients of the disease are severely afflicted with progressive neuro- and cardio-degeneration. The main cause of FA is due to the deficient expression of the mitochondrial protein, frataxin, and its deficiency has been well reported to be associated with oxidative stress and losses in energy metabolism. The major aim of this thesis was to elucidate the molecular mechanisms involved in the dysregulated anti-oxidative response in frataxin deficiency, which is responsible for the exacerbation of oxidative stress in FA. In light of the disease-associated deficits in mitochondrial bioenergetics and stress, this thesis then sought to explore the involvement of mitochondrial dynamics in the pathogenesis of FA. Considering these two pathological aspects of the disease, the thesis further assessed the efficacy of two different treatments aimed at restoring antioxidant defence and energy metabolism in vivo in frataxin deficiency. Results from these investigations are significant due to their potential applications and relevance to the development of future therapeutics for FA patients. This dissertation is comprised of 6 chapters: a comprehensive literature review (Chapter 1 Introduction); a materials and methods chapter (Chapter 2 Materials and Methods); 3 results chapters (Chapter 3 – 5); and a general discussion of principle findings and future directions chapter (Chapter 6 Discussion and Future Directions). viii Chapter 3: Various studies in models of FA have previously reported a decrease in the expression of the master regulator of antioxidant response, nuclear factor-erythroid 2-related factor-2 (Nrf2), due to unknown mechanisms. This chapter herein, examined the Nrf2-Keap1 signalling pathway using a mouse conditional frataxin knockout (KO) model of FA, by comparing the heart and skeletal muscle in these mice. The frataxin KO hearts exhibited fatal cardiomyopathy, while the skeletal muscle was asymptomatic. These two tissue-types demonstrated contrasting molecular alterations. In the KO heart, protein oxidation and decreased GSH:GSSG ratio were observed, as well as decreased total and nuclear Nrf2 expression and increased Keap1 levels. However, the skeletal muscle did not demonstrate these alterations. Notably, for the first time, a mechanism involving Gsk3β-signalling in the activation of nuclear Nrf2 export and/or degradation machinery was demonstrated in the KO heart. This process involved the increased activation of Gsk3β, increased Fyn phosphorylation, and the nuclear accumulation of β-TrCP to facilitate Nrf2 nuclear export. Furthermore, Nrf2-DNA-binding activity and the mRNA expression of Nrf2-targets were decreased in the frataxin KO mice. However, certain Nrf2 antioxidant targets, namely, NADPH quinone oxidoreductase-1 (Nqo1), glutathione-S-transferase-Mu1 (Gstm1), and thioredoxin reductase 1 (TxnRD1), demonstrated increased protein levels in the KO heart. In general, two potential mechanisms could be responsible for the reduced Nrf2 levels in the frataxin-deficient hearts: (1) increased cytosolic Keap1 levels, and (2) the activation of Gsk3β-signalling or the Gsk3β-Fyn axis that decreases nuclear Nrf2 levels. On the other hand, the frataxin-deficient skeletal muscle had no decrease in Nrf2 levels and had contrasting results to the heart. Collectively, these findings have revealed tissue-specific ix alterations in frataxin deficiency, but more importantly, the data have uncovered potential mechanisms that could significantly dysregulate the anti-oxidative response in FA. Chapter 4: The mitochondrion is an essential organelle that maintains cellular function and health through its role in energy production. The mitochondrion protects the cell from oxidative stress by maintaining its homeostasis with critically dynamic processes of mitochondrial biogenesis, fusion/fission, and mitophagy. An imbalance between oxidative stress formation and endogenous antioxidant processes can induce mitochondrial protein defects that can severely disrupt mitochondrial homeostasis. This can lead to mitochondrial dysfunction, which is accompanied by mitochondrial protein oxidation and mitochondrial DNA (mtDNA) damage, culminating in the depletion of ATP and NAD+, apoptosis and organ failure. The heart and the nervous systems, which have an abundance of mitochondria, are most vulnerable to mitochondrial protein dysfunction, as evident in a number of belligerent human disease states. FA is also regarded as a mitochondrial disease, due to the role of frataxin in mitochondrial functions. Frataxin deficiency leads to a defect in mitochondrial iron metabolism and oxidative stress that potentiates the pathology of the disease. However, alterations to mitochondrial homeostasis have not been fully elucidated in the pathogenesis of FA. Using the aforementioned frataxin KO mice model of FA that develops dilated cardiomyopathy, a number of key observations were found in the KO hearts relative to their wild-type littermates: (1) irregular mitochondrial morphologies and abnormal structure of cristae; (2) increased Parp activation, decreased NAD+:NADH ratio and reduced Sirt1 activity, (3) increased protein markers of mitochondrial biogenesis and dynamics (both fusion and fission), and (4) increased autophagic x flux at 10-weeks of age. These novel findings demonstrate significant changes to mitochondrial homeostasis in the condition of frataxin deficiency. Not only does this illustrate the importance of maintaining mitochondrial homeostasis in cardio-degenerative diseases, but it offers the potential for the development of new treatments that target mitochondrial function. Chapter 5: Results from Chapter 3 have found multiple molecular mechanisms involved in the dysregulation of the Nrf2 signalling pathway that negatively affects the anti-oxidative response in frataxin-deficient condition. Data from Chapter 4 have demonstrated significant alterations to mitochondrial morphologies, dynamics, and function in the frataxin KO mice with age. Taken together, these results indicate the critical involvement of oxidative stress, mitochondrial dysfunction and decreased bioenergetics in the pathogenesis of FA. Since there are currently limited treatments available for FA patients, there is an urgent need to develop new therapies that focuses on ameliorating these pathological deficits of the disease. This chapter herein examined the potential therapeutic effects of two agents, namely, N-acetylcysteine (NAC) in the supplementation of the antioxidant glutathione, and the novel compound, 6-methoxy-2-salicylaldehyde nicotinoyl hydrazine (SNH6), developed in our laboratory that has a dual-mechanism of action mediated by NAD+ supplementation and iron chelation. Using the previously described MCK mouse model of FA, these animals were treated from the asymptomatic age of 4-weeks-old, up until 9-weeks of age, where the animal displays an overt dilated cardiomyopathy. In general, iron deposits, interstitial fibrosis, and enlargement of cardiac muscle fibre size were observed in histological examinations of the KO hearts treated with either NAC or SNH6. Hence, the treatments did not attenuate disease progression or prevent the xi development of cardiac hypertrophy. Despite these observations, the treatment with SNH6 did significantly increase NAD+ levels, and as a result, there was increased sirtuin 1 and Parp activities in the SNH6-treated KO hearts. Hence, SNH6-supplementation of NAD+ potentially restored, in part, mitochondrial function and dynamics, despite that it did not increase the NAD+:NADH ratio and ATP levels. Collectively, increasing endogenous antioxidant levels and NAD+ supplementation are two different, but important therapeutic strategies that deserves further investigation. Particularly, the therapeutic use of the novel agent, SNH6, and the supplementation of NAD+, holds promise for the development of novel therapeutic strategies for FA patients. In conclusion, this dissertation has elucidated the molecular mechanisms involved in the dysregulation of anti-oxidative response and mitochondrial dynamics in the condition of frataxin deficiency of FA. The research in this thesis has enhanced our understanding of the pathophysiology of the disease and its associated cardiomyopathy, which offers new insights into the development of potential therapeutics. Thus, the significance of this dissertation is in its relevance to the advancement of future therapies for effective FA treatment.