Diagnostics for suspected mitochondrial disease (MD) can be challenging and necessitate invasive procedures like muscle biopsy. This is due to the extremely broad genetic and phenotypic spectrum, ...disease genes on both nuclear and mitochondrial DNA (mtDNA), and the tissue specificity of mtDNA variants. Exome sequencing (ES) has revolutionized the diagnostics for MD. However, the nuclear and mtDNA are investigated with separate tests, increasing costs and duration of diagnostics. The full potential of ES is often not exploited as the additional analysis of “off‐target reads” deriving from the mtDNA can be used to analyze both genomes. We performed mtDNA analysis by ES of 2111 cases in a clinical setting. We further assessed the recall rate and precision as well as the estimation of heteroplasmy by ES data by comparison with targeted mtDNA next generation sequencing in 49 cases. ES identified known pathogenic mtDNA point mutations in 38 individuals, increasing the diagnostic yield by nearly 2%. Analysis of mtDNA variants by ES had a high recall rate (96.2 ± 5.6%) and an excellent precision (99.5 ± 2.2%) when compared to the gold standard of targeted mtDNA next generation sequencing. ES estimated heteroplasmy levels with an average difference of 6.6 ± 3.8%, sufficient for clinical decision making. Taken together, the mtDNA analysis from ES is of sufficient quality for clinical diagnostics. We therefore propose ES, investigating both nuclear and mtDNA, as first line test in individuals with suspected MD. One should be aware, that a negative result does not exclude MD and necessitates further test (in additional tissues).
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FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Microglia adopt numerous fates with homeostatic microglia (HM) and a microglial neurodegenerative phenotype (MGnD) representing two opposite ends. A number of variants in genes selectively expressed ...in microglia are associated with an increased risk for neurodegenerative diseases such as Alzheimer's disease (AD) and frontotemporal lobar degeneration (FTLD). Among these genes are progranulin (GRN) and the triggering receptor expressed on myeloid cells 2 (TREM2). Both cause neurodegeneration by mechanisms involving loss of function. We have now isolated microglia from Grn−/− mice and compared their transcriptomes to those of Trem2−/− mice. Surprisingly, while loss of Trem2 enhances the expression of genes associated with a homeostatic state, microglia derived from Grn−/− mice showed a reciprocal activation of the MGnD molecular signature and suppression of gene characteristic for HM. The opposite mRNA expression profiles are associated with divergent functional phenotypes. Although loss of TREM2 and progranulin resulted in opposite activation states and functional phenotypes of microglia, FDG (fluoro‐2‐deoxy‐d‐glucose)‐μPET of brain revealed reduced glucose metabolism in both conditions, suggesting that opposite microglial phenotypes result in similar wide spread brain dysfunction.
Synopsis
Microglia from Grn−/− & Trem2−/− mice display opposite molecular signatures. While microglia are either locked in a hyperactivated or homeostatic state, Grn−/− & Trem2−/− mice both show reduced glucose metabolism, suggesting that opposite microglial phenotypes result in similar brain dysfunction.
First demonstration that microglia from both extremes of their functional stages cause brain wide dysfunctions.
This study indicates that the therapeutic window for microglial modulation is rather narrow and care must be taken to balance microglial activity.
Microglia from Grn−/− & Trem2−/− mice display opposite molecular signatures. While microglia are either locked in a hyperactivated or homeostatic state, Grn−/− & Trem2−/− mice both show reduced glucose metabolism, suggesting that opposite microglial phenotypes result in similar brain dysfunction.
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FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Alterations of Ca
homeostasis have been implicated in a wide range of neurodegenerative diseases. Ca
efflux from the endoplasmic reticulum into the cytoplasm is controlled by binding of inositol ...1,4,5-trisphosphate to its receptor. Activated inositol 1,4,5-trisphosphate receptors are then rapidly degraded by the endoplasmic reticulum-associated degradation pathway. Mutations in genes encoding the neuronal isoform of the inositol 1,4,5-trisphosphate receptor (ITPR1) and genes involved in inositol 1,4,5-trisphosphate receptor degradation (ERLIN1, ERLIN2) are known to cause hereditary spastic paraplegia (HSP) and cerebellar ataxia. We provide evidence that mutations in the ubiquitin E3 ligase gene RNF170, which targets inositol 1,4,5-trisphosphate receptors for degradation, are the likely cause of autosomal recessive HSP in four unrelated families and functionally evaluate the consequences of mutations in patient fibroblasts, mutant SH-SY5Y cells and by gene knockdown in zebrafish. Our findings highlight inositol 1,4,5-trisphosphate signaling as a candidate key pathway for hereditary spastic paraplegias and cerebellar ataxias and thus prioritize this pathway for therapeutic interventions.
Background
Congenital myasthenic syndromes (CMS) are a heterogeneous group of disorders caused by genetic defects resulting in impaired neuromuscular transmission. Although effective treatments are ...available, CMS is probably underdiagnosed, and systematic clinico-genetic investigations are warranted.
Methods
We used a nationwide approach to collect Austrian patients with genetically confirmed CMS. We provide a clinical and molecular characterization of this cohort and aimed to ascertain the current frequency of CMS in Austria.
Results
Twenty-eight cases with genetically confirmed CMS were identified, corresponding to an overall prevalence of 3.1 per million (95% CI 2.0–4.3) in Austria. The most frequent genetic etiology was
CHRNE
(
n
= 13), accounting for 46.4% of the cohort. Within this subgroup, the variant c.1327del, p.(Glu443Lysfs*64) was detected in nine individuals. Moreover, causative variants were found in
DOK7
(
n
= 4),
RAPSN
(
n
= 3),
COLQ
(
n
= 2),
GMPPB
(
n
= 2),
CHAT
(
n
= 1),
COL13A1
(
n
= 1),
MUSK
(
n
= 1) and
AGRN
(
n
= 1). Clinical onset within the first year of life was reported in one half of the patients. Across all subtypes, the most common symptoms were ptosis (85.7%), lower limb (67.9%), upper limb (60.7%) and facial weakness (60.7%). The majority of patients (96.4%) received specific treatment, including acetylcholinesterase inhibitors in 20, adrenergic agonists in 11 and 3,4-diaminopyridine in nine patients.
Conclusions
Our study presents the first systematic characterization of individuals with CMS in Austria, providing prevalence estimates and genotype–phenotype correlations that may help to improve the diagnostic approach and patient management.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Ferrodoxin reductase (FDXR) deficiency is a mitochondrial disease described in recent years primarily in association with optic atrophy, acoustic neuropathy, and developmental delays. Here, we ...identified seven unpublished patients with FDXR deficiency belonging to six independent families. These patients show a broad clinical spectrum ranging from Leigh syndrome with early demise and severe infantile‐onset encephalopathy, to milder movement disorders. In total nine individual pathogenic variants, of which seven were novel, were identified in FDXR using whole exome sequencing in suspected mitochondrial disease patients. Over 80% of these variants are missense, a challenging variant class in which to determine pathogenic consequence, especially in the setting of nonspecific phenotypes and in the absence of a reliable biomarker, necessitating functional validation. Here we implement an Arh1‐null yeast model to confirm the pathogenicity of variants of uncertain significance in FDXR, bypassing the requirement for patient‐derived material.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Neurodegeneration with brain iron accumulation (NBIA) are a group of clinically and genetically heterogeneous diseases characterized by iron overload in basal ganglia and progressive ...neurodegeneration. Little is known about the epidemiology of NBIA disorders. In the absence of large-scale population-based studies, obtaining reliable epidemiological data requires innovative approaches.
All pathogenic variants were collected from the 13 genes associated with autosomal recessive NBIA (PLA2G6, PANK2, COASY, ATP13A2, CP, AP4M1, FA2H, CRAT, SCP2, C19orf12, DCAF17, GTPBP2, REPS1). The allele frequencies of these disease-causing variants were assessed in exome/genome collections: the Genome Aggregation Database (gnomAD) and our in-house database. Lifetime risks were calculated from the sum of allele frequencies in the respective genes under assumption of Hardy-Weinberg equilibrium.
The combined estimated lifetime risk of all 13 investigated NBIA disorders is 0.88 (95% confidence interval 0.70–1.10) per 100,000 based on the global gnomAD dataset (n = 282,912 alleles), 0.92 (0.65–1.29) per 100,000 in the European gnomAD dataset (n = 129,206), and 0.90 (0.48–1.62) per 100,000 in our in-house database (n = 44,324). Individually, the highest lifetime risks (>0.15 per 100,000) are found for disorders caused by variants in PLA2G6, PANK2 and COASY.
This population-genetic estimation on lifetime risks of recessive NBIA disorders reveals frequencies far exceeding previous population-based numbers. Importantly, our approach represents lifetime risks from conception, thus including prenatal deaths. Understanding the true lifetime risk of NBIA disorders is important in estimating disease burden, allocating resources and targeting specific interventions.
This work was carried out in the framework of TIRCON (“Treat Iron-Related Childhood-Onset Neurodegeneration”).
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Pontocerebellar hypoplasia (PCH) describes a group of rare heterogeneous neurodegenerative diseases with prenatal onset. Here we describe eight children with PCH from four unrelated families ...harboring the homozygous MINPP1 (NM_004897.4) variants; c.75_94del, p.(Leu27Argfs*39), c.851 C > A, p.(Ala284Asp), c.1210 C > T, p.(Arg404*), and c.992 T > G, p.(Ile331Ser). The homozygous p.(Leu27Argfs*39) change is predicted to result in a complete absence of MINPP1. The p.(Arg404*) would likely lead to a nonsense mediated decay, or alternatively, a loss of several secondary structure elements impairing protein folding. The missense p.(Ala284Asp) affects a buried, hydrophobic residue within the globular domain. The introduction of aspartic acid is energetically highly unfavorable and therefore predicted to cause a significant reduction in protein stability. The missense p.(Ile331Ser) affects the tight hydrophobic interactions of the isoleucine by the disruption of the polar side chain of serine, destabilizing the structure of MINPP1. The overlap of the above-mentioned genotypes and phenotypes is highly improbable by chance. MINPP1 is the only enzyme that hydrolyses inositol phosphates in the endoplasmic reticulum lumen and several studies support its role in stress induced apoptosis. The pathomechanism explaining the disease mechanism remains unknown, however several others genes of the inositol phosphatase metabolism (e.g., INPP5K, FIG4, INPP5E, ITPR1) are correlated with phenotypes of neurodevelopmental disorders. Taken together, we present MINPP1 as a novel autosomal recessive pontocerebellar hypoplasia gene.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Motile and non-motile cilia play critical roles in mammalian development and health. These organelles are composed of a 1000 or more unique proteins, but their assembly depends entirely on proteins ...synthesized in the cell body and transported into the cilium by intraflagellar transport (IFT). In mammals, malfunction of non-motile cilia due to IFT dysfunction results in complex developmental phenotypes that affect most organs. In contrast, disruption of motile cilia function causes subfertility, disruption of the left-right body axis, and recurrent airway infections with progressive lung damage. In this work, we characterize allele specific phenotypes resulting from IFT74 dysfunction in human and mice. We identified two families carrying a deletion encompassing IFT74 exon 2, the first coding exon, resulting in a protein lacking the first 40 amino acids and two individuals carrying biallelic splice site mutations. Homozygous exon 2 deletion cases presented a ciliary chondrodysplasia with narrow thorax and progressive growth retardation along with a mucociliary clearance disorder phenotype with severely shorted cilia. Splice site variants resulted in a lethal skeletal chondrodysplasia phenotype. In mice, removal of the first 40 amino acids likewise results in a motile cilia phenotype but with little effect on primary cilia structure. Mice carrying this allele are born alive but are growth restricted and developed hydrocephaly in the first month of life. In contrast, a strong, likely null, allele of Ift74 in mouse completely blocks ciliary assembly and causes severe heart defects and midgestational lethality. In vitro studies suggest that the first 40 amino acids of IFT74 are dispensable for binding of other IFT subunits but are important for tubulin binding. Higher demands on tubulin transport in motile cilia compared to primary cilia resulting from increased mechanical stress and repair needs could account for the motile cilia phenotype observed in human and mice.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Mitochondrial disorders are a group of rare diseases, caused by nuclear or mitochondrial DNA mutations. Their marked clinical and genetic heterogeneity as well as referral and ascertainment biases ...render phenotype-based prevalence estimations difficult. Here we calculated the lifetime risk of all known autosomal recessive mitochondrial disorders on basis of genetic data.
We queried the publicly available Genome Aggregation Database (gnomAD) and our in-house exome database to assess the allele frequency of disease-causing variants in genes associated with autosomal recessive mitochondrial disorders. Based on this, we estimated the lifetime risk of 249 autosomal recessive mitochondrial disorders. Three of these disorders and phenylketonuria (PKU) served as a proof of concept since calculations could be aligned with known birth prevalence data from newborn screening reports.
The estimated lifetime risks are very close to newborn screening data (where available), supporting the validity of the approach. For example, calculated lifetime risk of PKU (16·0/100,000) correlates well with known birth prevalence data (18·7/100,000). The combined estimated lifetime risk of 249 investigated mitochondrial disorders is 31·8 (20·9–50·6)/100,000 in our in-house database, 48·4 (40·3–58·5)/100,000 in the European gnomAD dataset, and 31·1 (26·7–36·3)/100,000 in the global gnomAD dataset. The disorders with the highest lifetime risk (> 3 per 100,000) were, in all datasets, those caused by mutations in the SPG7, ACADM, POLG and SLC22A5 genes.
We provide a population-genetic estimation on the lifetime risk of an entire class of monogenic disorders. Our findings reveal the substantial cumulative prevalence of autosomal recessive mitochondrial disorders, far above previous estimates. These data will be very important for assigning diagnostic a priori probabilities, and for resource allocation in therapy development, public health management and biomedical research.
German Federal Ministry of Education and Research.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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