Answer to Gerber et al. "Autosomal recessive pathogenic MSTO1 variants in hereditary optic atrophy".

Gal et al address the issues raised by Gerber et al and reiterate that patients in their study showed decreased Misato homolog 1 (MSTO1) mRNA and protein levels, but also confirm finding of Gerber et al that the mutation is in MSTO2p pseudogene. Whether MSTO2p variant contributes to the observed decrease in MSTO1 levels in patients remains unclear.

Multilevel evidence of MECP2-associated mitochondrial dysfunction and its therapeutic implications.

We present a male patient carrying a pathogenic MECP2 p. Arg179Trp variant with predominant negative psychiatric features and multilevel evidence of mitochondrial dysfunction who responded to the cariprazine treatment. He had delayed speech development and later experienced severe social anxiety, learning disabilities, cognitive slowing, and predominant negative psychiatric symptoms associated with rigidity. Clinical examinations showed multisystemic involvement. Together with elevated ergometric lactate levels, the clinical picture suggested mitochondrial disease, which was also supported by muscle histopathology. Exploratory transcriptome analysis also revealed the involvement of metabolic and oxidative phosphorylation pathways. Whole-exome sequencing identified a pathogenic MECP2 variant, which can explain both the dopamine imbalance and mitochondrial dysfunction in this patient. Mitochondrial dysfunction was previously suggested in classical Rett syndrome, and we detected related phenotype evidence on multiple consistent levels for the first time in a MECP2 variant carrier male. This study further supports the importance of the MECP2 gene in the mitochondrial pathways, which can open the gate for more personalized therapeutic interventions. Good cariprazine response highlights the role of dopamine dysfunction in the complex psychiatric symptoms of Rett syndrome. This can help identify the optimal treatment strategy from a transdiagnostic perspective instead of a classical diagnostic category.

Genetic landscape of early-onset dementia in Hungary.

INTRODUCTION: Early-onset dementias (EOD) are predominantly genetically determined, but the underlying disease-causing alterations are often unknown. The most frequent forms of EODs are early-onset Alzheimer's disease (EOAD) and frontotemporal dementia (FTD). PATIENTS: This study included 120 Hungarian patients with EOD (48 familial and 72 sporadic) which had a diagnosis of EOAD (n = 49), FTD (n = 49), or atypical dementia (n = 22). RESULTS: Monogenic dementia was detected in 15.8% of the patients. A pathogenic hexanucleotide repeat expansion in the C9ORF72 gene was present in 6.7% of cases and disease-causing variants were detected in other known AD or FTD genes in 6.7% of cases (APP, PSEN1, PSEN2, GRN). A compound heterozygous alteration of the TREM2 gene was identified in one patient and heterozygous damaging variants in the CSF1R and PRNP genes were detected in two other cases. In two patients, the coexistence of several heterozygous damaging rare variants associated with neurodegeneration was detected (1.7%). The APOE genotype had a high odds ratio for both the APOE ɛ4/3 and the ɛ4/4 genotype (OR = 2.7 (95%CI = 1.3-5.9) and OR = 6.5 (95%CI = 1.4-29.2), respectively). In TREM2, SORL1, and ABCA7 genes, 5 different rare damaging variants were detected as genetic risk factors. These alterations were not present in the control group. CONCLUSION: Based on our observations, a comprehensive, targeted panel of next-generation sequencing (NGS) testing investigating several neurodegeneration-associated genes may accelerate the path to achieve the proper genetic diagnosis since phenotypes are present on a spectrum. This can also reveal hidden correlations and overlaps in neurodegenerative diseases that would remain concealed in separated genetic testing.

Correlation of GAA Genotype and Acid-α-Glucosidase Enzyme Activity in Hungarian Patients with Pompe Disease.

Pompe disease is caused by the accumulation of glycogen in the lysosomes due to a deficiency of the lysosomal acid-α-glucosidase (GAA) enzyme. Depending on residual enzyme activity, the disease manifests two distinct phenotypes. In this study, we assess an enzymatic and genetic analysis of Hungarian patients with Pompe disease. Twenty-four patients diagnosed with Pompe disease were included. Enzyme activity of acid-α-glucosidase was measured by mass spectrometry. Sanger sequencing and an MLPA of the GAA gene were performed in all patients. Twenty (83.33%) patients were classified as having late-onset Pompe disease and four (16.66%) had infantile-onset Pompe disease. Fifteen different pathogenic GAA variants were detected. The most common finding was the c.-32-13 T > G splice site alteration. Comparing the α-glucosidase enzyme activity of homozygous cases to the compound heterozygous cases of the c.-32-13 T > G disease-causing variant, the mean GAA activity in homozygous cases was significantly higher. The lowest enzyme activity was found in cases where the c.-32-13 T > G variant was not present. The localization of the identified sequence variations in regions encoding the crucial protein domains of GAA correlates with severe effects on enzyme activity. A better understanding of the impact of pathogenic gene variations may help earlier initiation of enzyme replacement therapy (ERT) if subtle symptoms occur. Further information on the effect of GAA gene variation on the efficacy of treatment and the extent of immune response to ERT would be of importance for optimal disease management and designing effective treatment plans.

New Insights of Phospholipase A2 Associated Neurodegeneration Phenotype Based on the Long-Term Follow-Up of a Large Hungarian Family.

INTRODUCTION: Phospholipase A2-associated Neurodegeneration (PLAN) is a group of neurodegenerative diseases associated with the alterations of PLA2G6. Some phenotype-genotype association are well known but there is no clear explanation why some cases can be classified into distinct subgroups, while others follow a continuous clinical spectrum. METHODS: Long-term neurological, and psychiatric follow-up, neuropathological, radiological, and genetic examinations, were performed in three affected girls and their family. RESULTS: Two 24-years old twins and their 22-years old sister harbored the p.P622S, and p.R600W mutation in PLA2G6. The age of onset and the most prominent presenting symptoms (gaze palsy, ataxia, dystonia, psychomotor regression indicated atypical neuroaxonal dystrophy (ANAD), however, optic atrophy, severe tetraparesis would fit into infantile neuroaxonal dystrophy (INAD). All siblings had hyperintensity in the globi pallidi and substantiae nigrae which is reported in ANAD, whereas it is considered a later neuroradiological marker in INAD. The slow progression, rigidity, bradykinesis, and the prominent psychiatric symptoms indicate PLA2G6-related dystonia-parkinsonism. Abnormal mitochondria, lipid accumulation and axonal spheroids were observed in the muscle and nerve tissue. Brain deposition appeared 6 years following the initial cerebellar atrophy. Mild MRI alterations were detected in the asymptomatic carrier parents. CONCLUSION: The colorful clinical symptoms, the slightly discordant phenotype, and the neuroimaging data in the family supports the view that despite the distinct definition of age-related phenotypes in PLAN, these are not strict disease categories, but rather a continuous phenotypic spectrum. The mild MRI alterations of the parents and the family history suggest that even heterozygous pathogenic variants might be associated with clinical symptoms, although systematic study is needed to prove this.

Novel dominant MPAN family with a complex genetic architecture as a basis for phenotypic variability.

OBJECTIVE: Our aim was to study a Hungarian family with autosomal dominantly inherited neurodegeneration with brain iron accumulation (NBIA) with markedly different intrafamilial expressivity. METHODS: Targeted sequencing and multiplex ligation-dependent probe amplification (MLPA) of known NBIA-associated genes were performed in many affected and unaffected members of the family. In addition, a trio whole-genome sequencing was performed to find a potential explanation of phenotypic variability. Neuropathologic analysis was performed in a single affected family member. RESULTS: The clinical phenotype was characterized by 3 different syndromes-1 with rapidly progressive dystonia-parkinsonism with cognitive deterioration, 1 with mild parkinsonism associated with dementia, and 1 with predominantly psychiatric symptoms along with movement disorder. A heterozygous stop-gain variation in the C19Orf12 gene segregated with the phenotype. Targeted sequencing of all known NBIA genes, and MLPA of PLA2G6 and PANK2 genes, as well as whole-genome sequencing in a trio from the family, revealed a unique constellation of oligogenic burden in 3 NBIA-associated genes (C19Orf12 p.Trp112Ter, CP p.Val105PhefsTer5, and PLA2G6 dup(ex14)). Neuropathologic analysis of a single case (39-year-old man) showed a complex pattern of alpha-synucleinopathy and tauopathy, both involving subcortical and cortical areas and the hippocampus. CONCLUSIONS: Our study expands the number of cases reported with autosomal dominant mitochondrial membrane protein-associated neurodegeneration and emphasizes the complexity of the genetic architecture, which might contribute to intrafamilial phenotypic variability.

Wernicke-Korsakoff syndrome associated with mtDNA disease.

INTRODUCTION: Wernicke encephalopathy (WE) and Wernicke-Korsakoff syndrome (WKS) are well-known disorders caused by thiamine deficiency. In addition to the classical concept of these diseases, some literature data suggest a connection between mitochondrial dysfunction and WE/WKS. Psychotic disorders and WKS seem to run in families, as the deficiency of the oxidative phosphorylation can be a trigger factor in psychotic events and WE/WKS as well. We present a patient harbouring the m.A3243G mtDNA mutation with the clinical and magnetic resonance imaging (MRI) findings of WKS who developed schizophrenia with predominantly negative symptoms some years later. CASE PRESENTATION: A 27-year-old woman was referred to our clinic with severe weight loss after severe vomiting episodes, memory dysfunction and gait ataxia. Family history, as well as clinical, imaging and laboratory findings suggested a mitochondrial aetiology of her symptoms. Brain MRI detected bilateral mild thalamic lesions and loss of corpus mammillae, indicating Wernicke encephalopathy. Genetic testing detected an m.A3243G mtDNA mutation, which has been frequently associated with mitochondrial encephalopathy with lactic acidosis and stroke-like episodes. High-dose vitamin B1 supplementation with supportive antioxidant therapy improved the patient's memory and learning disturbance; however, some months later she developed psychosis with predominantly negative symptoms and her cognitive functions deteriorated again. Both cognitive and negative symptoms responded well to cariprazine monotherapy. DISCUSSION: Mitochondrial disease due to mtDNA alteration can be a rare cause of WE. In addition to vitamin B1 supplementation, cariprazine with significant dopamine D3 receptor binding can be useful to treat the predominantly negative symptoms and cognitive dysfunction in patients with mitochondrial dysfunction. CONCLUSION: We assume that patients with a mitochondrial disorder might be prone to develop WE/WKS and therefore need tailored supportive therapy during metabolic crisis as well as symptom-based personalized antipsychotic treatment.

Hereditary Parkinson's disease as a new clinical manifestation of the damaged POLG gene / [Az örökletes Parkinson-kór mint a POLG-gén károsodásának új klinikai megjelenési formája]

The protein product of the nuclear-encoded POLG gene plays a key role in the maintenance of mitochondrial DNA replication, and its failure causes multi-system diseases with varying severity. The clinical spectrum is extremely wide, and the most common symptoms include ptosis, myoclonus, epilepsy, myopathy, sensory ataxia, parkinsonism, cognitive decline and infertility. Now, it is known that mitochondrial dysfunction in Parkinson's disease plays a key role in the loss of dopaminergic neurons in the substantia nigra. Therefore, changes in the POLG gene may influence the development of various hereditary neurodegenerative diseases, including monogenic parkinsonism. However, only limited information is available on the relationship between Parkinson's disease and POLG gene and until now, there are no available data about the Hungarian population. In our study, we performed a next-generation sequencing study of 67 Hungarian patients with parkinsonism and analyzed the potentially damaging alterations in the POLG gene. 3 patients have been identified with a potential pathogen variant. In this study, we would like to call attention to the fact that during the differential diagnosis of parkinsonism, the possible involvement of POLG gene should be kept in mind. Especially in the presence of additional symptoms, such as ophthalmoparesis, non-vascular white matter lesions, psychiatric comorbidity, and relatively early age of onset, the POLG gene should be taken into consideration. Based on previous data from the literature and our own experience, we have summarized a possible diagnostic approach for POLG-associated parkinsonism. Orv Hetil. 2020; 161(20): 821-828.

Broadening the phenotype of the TWNK gene associated Perrault syndrome.

BACKGROUND: Perrault syndrome is a genetically heterogenous, very rare disease, characterized clinically by sensorineural hearing loss, ovarian dysfunction and neurological symptoms. We present the case of a 33 years old female patient with TWNK-associated Perrault syndrome. The TWNK gene is coding the mitochondrial protein Twinkle and currently there are only two reports characterizing the phenotype of TWNK-associated Perrault syndrome. None of these publications reported about special brain MRI alterations and neuropathological changes in the muscle and peripheral nerves. CASE PRESENTATION: Our patients with TWNK-dependent Perrault syndrome had severe bilateral hypoacusis, severe ataxia, polyneuropathy, lower limb spastic paraparesis with pyramidal signs, and gonadal dysgenesis. Psychiatric symptoms such as depression and paranoia were present as well. Brain MRI observed progressive cerebellar hyperintensive signs associated with cerebellar, medulla oblongata and cervical spinal cord atrophy. Light microscopy of the muscle biopsy detected severe neurogenic lesions. COX staining was centrally reduced in many muscle fibers. Both muscle and sural nerve electron microscopy detected slightly enlarged mitochondria with abnormal cristae surrounded by lipid vacuoles. In the sural nerve, dystrophic axons had focally uncompacted myelin lamellae present. Genetic investigation revealed multiple mtDNA deletion and compound heterozygous mutations of the TWNK gene (c.1196 A > G, c.1358 G > A). CONCLUSION: This study demonstrates that TWNK associated Perrault syndrome has a much broader phenotype as originally published. The coexistence of severe hypoacusis, spastic limb weakness, ataxia, polyneuropathy, gonadal dysgensia, hyperintense signals in the cerebellum and the presence of the mtDNA multiple deletion could indicate the impairment of the TWNK gene. This is the first report about pyramidal tract involvement and cerebellar MRI alteration associated with TWNK-related Perrault syndrome.

The Role of Genetic Testing in the Clinical Practice and Research of Early-Onset Parkinsonian Disorders in a Hungarian Cohort: Increasing Challenge in Genetic Counselling, Improving Chances in Stratification for Clinical Trials.

The genetic analysis of early-onset Parkinsonian disorder (EOPD) is part of the clinical diagnostics. Several genes have been implicated in the genetic background of Parkinsonism, which is clinically indistinguishable from idiopathic Parkinson's disease. The identification of patient's genotype could support clinical decision-making process and also track and analyse outcomes in a comprehensive fashion. The aim of our study was to analyse the genetic background of EOPD in a Hungarian cohort and to evaluate the clinical usefulness of different genetic investigations. The age of onset was between 25 and 50 years. To identify genetic alterations, multiplex ligation-dependent probe amplification (n = 142), Sanger sequencing of the most common PD-associated genes (n = 142), and next-generation sequencing (n = 54) of 127 genes which were previously associated to neurodegenerative disorders were carried out. The genetic analysis identified several heterozygous damaging substitutions in PD-associated genes (C19orf12, DNAJC6, DNAJC13, EIF4G1, LRRK2, PRKN, PINK1, PLA2G6, SYNJ1). CNVs in PRKN and SNCA genes were found in five patients. In our cohort, nine previously published genetic risk factors were detected in three genes (GBA, LRRK2, and PINK1). In nine cases, two or three coexisting pathogenic mutations and risk variants were identified. Advances of sequencing technologies make it possible to aid diagnostics of PD by widening the scope of analysis to genes which were previously linked to other neurodegenerative disorders. Our data suggested that rare damaging variants are enriched versus neutral variants, among PD patients in the Hungarian population, which raise the possibility of an oligogenic effect. Heterozygous mutations of multiple recessive genes involved in the same pathway may perturb the molecular process linked to PD pathogenesis. Comprehensive genetic assessment of individual patients can rarely reveal monogenic cause in EOPD, although it may identify the involvement of multiple PD-associated genes in the background of the disease and may facilitate the better understanding of clinically distinct phenocopies. Due to the genetic complexity of the disease, genetic counselling and management is getting more challenging. Clinical geneticist should be prepared for counselling of patients with coexisting disease-causing mutations and susceptibility factors. At the same time, genomic-based stratification has increasing importance in future clinical trials.

Comprehensive Analysis of Rare Variants of 101 Autism-Linked Genes in a Hungarian Cohort of Autism Spectrum Disorder Patients.

BACKGROUND: Autism spectrum disorder (ASD) is genetically and phenotypically heterogeneous. Former genetic studies suggested that both common and rare genetic variants play a role in the etiology. In this study, we aimed to analyze rare variants detected by next generation sequencing (NGS) in an autism cohort from Hungary. METHODS: We investigated the yield of NGS panel sequencing of an unselected ASD cohort (N = 174 ) for the detection of ASD associated syndromes. Besides, we analyzed rare variants in a common disease-rare variant framework and performed rare variant burden analysis and gene enrichment analysis in phenotype based clusters. RESULTS: We have diagnosed 13 molecularly proven syndromic autism cases. Strongest indicators of syndromic autism were intellectual disability, epilepsy or other neurological plus symptoms. Rare variant analysis on a cohort level confirmed the association of five genes with autism (AUTS2, NHS, NSD1, SLC9A9, and VPS13). We found no correlation between rare variant burden and number of minor malformation or autism severity. We identified four phenotypic clusters, but no specific gene was enriched in a given cluster. CONCLUSION: Our study indicates that NGS panel gene sequencing can be useful, where the clinical picture suggests a clinically defined syndromic autism. In this group, targeted panel sequencing may provide reasonable diagnostic yield. Unselected NGS panel screening in the clinic remains controversial, because of uncertain utility, and difficulties of the variant interpretation. However, the detected rare variants may still significantly influence autism risk and subphenotypes in a polygenic model, but to detect the effects of these variants larger cohorts are needed.

[Etiological factors of sensorineural hearing loss in children after cochlear implantation]. / Halláscsökkenést okozó etiológiai tényezok cochlearis implantáción átesett gyermekekben.

Introduction: Congenital sensorineural hearing loss is one of the most common sensory defects affecting 1-3 children per 1000 newborns. There are a lot of causes which result in congenital hearing loss, the most common is the genetic origin, but infection, cochlear malformation or other acquired causes can be reasons as well. Aim: The aim of this study was to establish the etiological factors of congenital profound sensorineural hearing loss in children who underwent cochlear implantation. Results: Our results show that the origin of the hearing loss was discovered in 62.9% of our patients. The most common etiological factor was the c.35delG mutation of the gap junction protein ß-2 gene, the allele frequency was 38.7% in our cohort. Infection constituted to 10.1%, and meningitis and cytomegalovirus infection were the second most common cause. 79.9% of our patients received sufficient hearing rehabilitation before the end of the speech development's period (6 years old), but 11.2% of our cases were still diagnosed late. Conclusions: Based on our data we can state that genetic evaluation is crucial in the diagnostic process of congenital profound sensorineural hearing loss. Sufficient hearing rehabilitation affects the whole life of the child, and by late cochlear implantation the speech development falls behind. We can decrease the ratio of the late implantation with the new protocol of newborn hearing screening, and with sufficient information provided to the colleagues, so the children may be referred to the proper center for rehabilitation without delay. Orv Hetil. 2019; 160(21): 822-828.

Analysis of GJB2 mutations and the clinical manifestation in a large Hungarian cohort.

PURPOSE: Pathogenic variants of the gap junction beta 2 (GJB2) gene are responsible for about 50% of hereditary non-syndromic sensorineural hearing loss (NSHL). In this study, we report mutation frequency and phenotype comparison of different GJB2 gene alterations in Hungarian NSHL patients. METHODS: The total coding region of the GJB2 gene was analyzed with Sanger or NGS sequencing for 239 patients with NSHL and 160 controls. RESULTS: Homozygous and compound heterozygous GJB2 mutations were associated with early onset serious clinical phenotype in 28 patients. In 24 patients, two deletion or nonsense mutations were detected in individuals with mainly prelingual NSHL. In compound heterozygous cases, a combination of deletion and missense mutations associated with milder postlingual NSHL. A further 25 cases harbored single heterozygous GJB2 mutations mainly associated with later onset, milder clinical phenotype. The most common mutation was the c.35delG deletion, with 12.6% allele frequency. The hearing loss was more severe in the prelingual groups. CONCLUSION: The mutation frequency of GJB2 in the investigated cohort is lower than in other European cohorts. The most serious cases were associated with homozygous and compound heterozygous mutations. In our cohort the hearing impairment and age of onset was not altered between in cases with only one heterozygous GJB2 mutation and wild type genotype, which may exclude the possibility of autosomal dominant inheritance. In early onset, severe to profound hearing loss cases, if the GJB2 analysis results in only one heterozygous alteration further next generation sequencing is highly recommended.

[Significance of whole exome sequencing in the diagnostics of rare neurological diseases - own experiences through a case presenting with ataxia]. / A teljesexom-szekvenálás jelentosége a ritka neurológiai betegségek diagnosztikájában ­ saját tapasztalatok egy ataxiás eset kapcsán.

Next generation sequencing (NGS) technologies reshape the diagnostics of rare neurological diseases. In the background of certain neurological symptoms, such as ataxia, many acquired and genetic causes may be present. Variations in a given gene can present with variable phenotypes, too. Because of this phenomenon, the conventional one gene sequencing approach often fails to identify the genetic background of a disease. Next generation sequencing panels allow to sequence 50-100 genes simultaneously, and if the disease stratification is not possible based on the clinical symptoms, whole exome sequencing can help in the diagnostic of genetic disorders with atypical presentation. This case study is about the exome sequencing of a patient with cerebellar ataxia. Genetic investigations identified rare variants in the SPG11 gene in association with the clinical phenotype, which gene was originally described in the background of hereditary spastic paraparesis. Our article highlights that in certain cases the variability of the leading presenting symptom makes it hard to select the correct gene panel. In our case the variants in the gene, formerly associated to hereditary spastic paraparesis, resulted in cerebellar ataxia initially, so even an ataxia NGS gene panel would not detect those. Orv Hetil. 2018; 159(28): 1163-1169.

Implementation of personalized medicine in Central-Eastern Europe: pitfalls and potentials based on citizen's attitude.

OBJECTIVE: Next-generation sequencing is increasingly utilized worldwide as a research and diagnostic tool and is anticipated to be implemented into everyday clinical practice. Since Central-Eastern European attitude toward genetic testing, especially broad genetic testing, is not well known, we performed a survey on this issue among Hungarian participants. METHODS: A self-administered questionnaire was distributed among patients and patient relatives at our neurogenetic outpatient clinic. Members of the general population were also recruited via public media. We used chi-square testing and binary logistic regression to examine factors influencing attitude. RESULTS: We identified a mixed attitude toward genetic testing. Access to physician consultation positively influenced attitude. A higher self-determined genetic familiarity score associated with higher perceived genetic influence score, which in turn associated with greater willingness to participate in genetic testing. Medical professionals constituted a skeptical group. CONCLUSIONS: We think that given the controversies and complexities of the next-generation sequencing field, the optimal clinical translation of NGS data should be performed in institutions which have the unique capability to provide interprofessional health education, transformative biomedical research, and crucial patient care. With optimization of the clinical translational process, improvement of genetic literacy may increase patient engagement and empowerment. RELEVANCE OF THE ARTICLE FOR PREDICTIVE PREVENTIVE AND PERSONALIZED MEDICINE: The paper highlights that in countries with relatively low-genetic literacy, a special strategy is needed to enhance the implementation of personalized medicine.

Genotypic and phenotypic spectrum of the most common causative genes of Charcot-Marie-Tooth disease in Hungarian patients.

Charcot-Marie-Tooth neuropathy (CMT) is a genetically and clinically heterogeneous group of neuromuscular disorders with an overall prevalence of 1 per 2500. Here we report the first comprehensive genetic epidemiology study of Hungarian CMT patients. 409 CMT1 and 122 CMT2 patients were enrolled and genetic testing of PMP22, GJB1, MPZ, EGR2 and MFN2 genes were performed routinely. NDRG1 and CTDP1 genes were screened only for founder mutations in Roma patients. Causative genetic mutations were identified in 67.2% of the CMT1 and in 33.6% of the CMT2 cases, which indicates an overall success rate of 59.9% in the study population. Considering all affected individuals, alterations were most frequently found in PMP22 (40.5%), followed by GJB1 (9.2%), MPZ (4.5%), MFN2 (2.5%), NDRG1 (1.5%), EGR2 (0.8%) and CTDP1 (0.8%). The phenotypic spectrum and the disease severity of the studied patients also varied broadly. Deafness and autoimmune disorders were more often associated with PMP22 duplication, while MFN2 and GJB1 mutations were frequently present with central nervous system abnormalities. Our study may be helpful in determining the strategy of genetic diagnostics in Hungarian CMT patients.

Recessive mutations in MSTO1 cause mitochondrial dynamics impairment, leading to myopathy and ataxia.

We report here the first families carrying recessive variants in the MSTO1 gene: compound heterozygous mutations were identified in two sisters and in an unrelated singleton case, who presented a multisystem complex phenotype mainly characterized by myopathy and cerebellar ataxia. Human MSTO1 is a poorly studied protein, suggested to have mitochondrial localization and to regulate morphology and distribution of mitochondria. As for other mutations affecting genes involved in mitochondrial dynamics, no biochemical defects typical of mitochondrial disorders were reported. Studies in patients' fibroblasts revealed that MSTO1 protein levels were strongly reduced, the mitochondrial network was fragmented, and the fusion events among mitochondria were decreased, confirming the deleterious effect of the identified variants and the role of MSTO1 in modulating mitochondrial dynamics. We also found that MSTO1 is mainly a cytosolic protein. These findings indicate recessive mutations in MSTO1 as a new cause for inherited neuromuscular disorders with multisystem features.

MSTO1 is a cytoplasmic pro-mitochondrial fusion protein, whose mutation induces myopathy and ataxia in humans.

The protein MSTO1 has been localized to mitochondria and linked to mitochondrial morphology, but its specific role has remained unclear. We identified a c.22G > A (p.Val8Met) mutation of MSTO1 in patients with minor physical abnormalities, myopathy, ataxia, and neurodevelopmental impairments. Lactate stress test and myopathological results suggest mitochondrial dysfunction. In patient fibroblasts, MSTO1 mRNA and protein abundance are decreased, mitochondria display fragmentation, aggregation, and decreased network continuity and fusion activity. These characteristics can be reversed by genetic rescue. Short-term silencing of MSTO1 in HeLa cells reproduced the impairment of mitochondrial morphology and dynamics observed in the fibroblasts without damaging bioenergetics. At variance with a previous report, we find MSTO1 to be localized in the cytoplasmic area with limited colocalization with mitochondria. MSTO1 interacts with the fusion machinery as a soluble factor at the cytoplasm-mitochondrial outer membrane interface. After plasma membrane permeabilization, MSTO1 is released from the cells. Thus, an MSTO1 loss-of-function mutation is associated with a human disorder showing mitochondrial involvement. MSTO1 likely has a physiologically relevant role in mitochondrial morphogenesis by supporting mitochondrial fusion.

Natural and Induced Mitochondrial Phosphate Carrier Loss: DIFFERENTIAL DEPENDENCE OF MITOCHONDRIAL METABOLISM AND DYNAMICS AND CELL SURVIVAL ON THE EXTENT OF DEPLETION.

The relevance of mitochondrial phosphate carrier (PiC), encoded by SLC25A3, in bioenergetics is well accepted. However, little is known about the mechanisms mediating the cellular impairments induced by pathological SLC25A3 variants. To this end, we investigated the pathogenicity of a novel compound heterozygous mutation in SLC25A3 First, each variant was modeled in yeast, revealing that substituting GSSAS for QIP within the fifth matrix loop is incompatible with survival on non-fermentable substrate, whereas the L200W variant is functionally neutral. Next, using skin fibroblasts from an individual expressing these variants and HeLa cells with varying degrees of PiC depletion, PiC loss of ∼60% was still compatible with uncompromised maximal oxidative phosphorylation (oxphos), whereas lower maximal oxphos was evident at ∼85% PiC depletion. Furthermore, intact mutant fibroblasts displayed suppressed mitochondrial bioenergetics consistent with a lower substrate availability rather than phosphate limitation. This was accompanied by slowed proliferation in glucose-replete medium; however, proliferation ceased when only mitochondrial substrate was provided. Both mutant fibroblasts and HeLa cells with 60% PiC loss showed a less interconnected mitochondrial network and a mitochondrial fusion defect that is not explained by altered abundance of OPA1 or MFN1/2 or relative amount of different OPA1 forms. Altogether these results indicate that PiC depletion may need to be profound (>85%) to substantially affect maximal oxphos and that pathogenesis associated with PiC depletion or loss of function may be independent of phosphate limitation when ATP requirements are not high.