Objective
Oculopharyngodistal myopathy (OPDM) is an adult‐onset neuromuscular disease characterized by progressive ptosis, dysarthria, ophthalmoplegia, and distal muscle weakness. Recent studies ...revealed that GGC repeat expansions in 5′‐UTR of LRP12, GIPC1, and NOTCH2NLC are associated with OPDM. Despite these advances, approximately 30% of OPDM patients remain genetically undiagnosed. Herein, we aim to investigate the genetic basis for undiagnosed OPDM patients in two unrelated Chinese Han families.
Methods
Parametric linkage analysis was performed. Long‐read sequencing followed by repeat‐primed polymerase chain reaction and amplicon length polymerase chain reaction were used to determine the genetic cause. Targeted methylation sequencing was implemented to detect epigenetic changes. The possible pathogenesis mechanism was investigated by quantitative polymerase chain reaction, immunoblotting, RNA fluorescence in situ hybridization, and immunofluorescence staining of muscle biopsy samples.
Results
The disease locus was mapped to 12q24.3. Subsequently, GGC repeat expansion in the promoter region of RILPL1 was identified in six OPDM patients from two families, findings consistent with a founder effect, designated as OPDM type 4. Targeted methylation sequencing revealed hypermethylation at the RILPL1 locus in unaffected individuals with ultralong expansion. Analysis of muscle samples showed no significant differences in RILPL1 mRNA or RILPL1 protein levels between patients and controls. Public CAGE‐seq data indicated that alternative transcription start sites exist upstream of the RefSeq‐annotated RILPL1 transcription start site. Strand‐specific RNA‐seq data revealed bidirectional transcription from the RILPL1 locus. Finally, fluorescence in situ hybridization/immunofluorescence staining showed that both sense and antisense transcripts formed RNA foci, and were co‐localized with hnRNPA2B1 and p62 in the intranuclear inclusions of OPDM type 4 patients.
Interpretation
Our findings implicate abnormal GGC repeat expansions in the promoter region of RILPL1 as a novel genetic cause for OPDM, and suggest a methylation mechanism and a potential RNA toxicity mechanism are involved in OPDM type 4 pathogenesis. ANN NEUROL 2022;92:512–526
Objective
Despite the increasing number of genes associated with Charcot–Marie‐Tooth (CMT) disease, many patients currently still lack appropriate genetic diagnosis for this disease. Autosomal ...dominant mutations in aminoacyl‐tRNA synthetases (ARSs) have been implicated in CMT. Here, we describe causal missense mutations in the gene encoding seryl‐tRNA synthetase 1 (SerRS) for 3 families affected with CMT.
Methods
Whole‐exome sequencing was performed in 16 patients and 14 unaffected members of 3 unrelated families. The functional impact of the genetic variants identified was investigated using bioinformatic prediction tools and confirmed using cellular and biochemical assays.
Results
Combined linkage analysis for the 3 families revealed significant linkage (Zmax LOD = 6.9) between the genomic co‐ordinates on chromosome 1: 108681600–110300504. Within the linkage region, heterozygous SerRS missense variants segregated with the clinical phenotype in the 3 families. The mutant SerRS proteins exhibited reduced aminoacylation activity and abnormal SerRS dimerization, which suggests the impairment of total protein synthesis and induction of eIF2α phosphorylation.
Interpretation
Our findings suggest the heterozygous SerRS variants identified represent a novel cause for autosomal dominant CMT. Mutant SerRS proteins are known to impact various molecular and cellular functions. Our findings provide significant advances on the current understanding of the molecular mechanisms associated with ARS‐related CMT. ANN NEUROL 2023;93:244–256
Primary familial brain calcification (PFBC) is a rare neurological disorder. Mutations in five genes (SLC20A2, PDGFRB, PDGFB, XPR1, and MYORG) have been linked to PFBC. Here, we used SYBR green‐based ...real‐time quantitative polymerase chain reaction (PCR) assay and denaturing high‐performance liquid chromatography analysis to detect copy number variants (CNVs) in 20 unrelated patients with PFBC, negatively sequenced for the five known genes. We identified three deletions in SLC20A2, including a large de novo full gene deletion and two exonic deletions confined to exon 2 and exon 6, respectively. Subsequent linked‐read whole‐genome sequencing of the patient with the large deletion showed a 1.7 Mb heterozygous deletion which removed the entire coding regions of SLC20A2 as well as 21 other genes. In the family with a deletion of exon 6, a missense variant of uncertain significance (SLC20A2: p.E267Q) also co‐segregated with the disease. Functional assay showed the deletion could result in significantly impaired phosphate transport, whereas the p.E267Q variant did not. Our results confirm that deletion in SLC20A2 is a causal mechanism for PFBC and highlight the importance of functional study for classifying a rare missense variant as (likely) pathogenic.
SLC20A2 deletions were identified in three of the 20 families with genetically unexplained primary familial brain calcification (PFBC), highlighting that copy number variants analysis should be considered, along with direct sequencing, in the genetic evaluation of PFBC patients.
Primary familial brain calcification (PFBC) is a progressive neurological disorder manifesting as bilateral brain calcifications in CT scan with symptoms as parkinsonism, dystonia, ataxia, ...psychiatric symptoms, etc. Recently, pathogenic variants in
MYORG
have been linked to autosomal recessive PFBC. This study aims to elucidate the mutational and clinical spectrum of
MYORG
mutations in a large cohort of Chinese PFBC patients with possible autosomal recessive or absent family history. Mutational analyses of
MYORG
were performed by Sanger sequencing in a cohort of 245 PFBC patients including 21 subjects from 10 families compatible with a possibly autosomal-recessive trait and 224 apparently sporadic cases. In-depth phenotyping and neuroimaging features were investigated in all patients with novel
MYORG
variants. Two nonsense variants (c.442C > T, p. Q148*; c.972C > A, p. Y324*) and two missense variants (c.1969G>C, p. G657R; c.2033C > G, p. P678R) of
MYORG
were identified in four sporadic PFBC patients, respectively. These four novel variants were absent in gnomAD, and their amino acid were highly conserved, suggesting these variants have a pathogenic impact. Patients with
MYORG
variants tend to display a homogeneous clinical spectrum, showing extensive brain calcification and parkinsonism, dysarthria, ataxia, or vertigo. Our findings supported the pathogenic role of
MYORG
variants in PFBC and identified two pathogenic variants (c.442C > T, c.972C > A), one likely pathogenic variant (c.2033C > G), and one variant of uncertain significance (c.1969G>C), further expanding the genetic and phenotypic spectrum of PFBC-
MYORG
.
Abstract
Primary familial brain calcification (PFBC) is a neurogenetic disorder characterized by bilateral calcified deposits in the brain. We previously identified that
MYORG
as the first pathogenic ...gene for autosomal recessive PFBC, and established a
Myorg
-KO mouse model. However,
Myorg
-KO mice developed brain calcifications until nine months of age, which limits their utility as a facile PFBC model system. Hence, whether there is another typical animal model for mimicking PFBC phenotypes in an early stage still remained unknown. In this study, we profiled the mRNA expression pattern of
myorg
in zebrafish, and used a morpholino-mediated blocking strategy to knockdown
myorg
mRNA at splicing and translation initiation levels. We observed multiple calcifications throughout the brain by calcein staining at 2–4 days post-fertilization in
myorg-
deficient zebrafish, and rescued the calcification phenotype by replenishing
myorg
cDNA. Overall, we built a novel model for PFBC via knockdown of
myorg
by antisense oligonucleotides in zebrafish, which could shorten the observation period and replenish the
Myorg
-KO mouse model phenotype in mechanistic and therapeutic studies.
IntroductionHereditary spastic paraplegias (HSPs) are uncommon but not rare neurodegenerative diseases. More than 100 pathogenic genes and loci related to spastic paraplegia symptoms have been ...reported. HSPs have the same core clinical features, including progressive spasticity in the lower limbs, though HSPs are heterogeneous (eg, clinical signs, MRI features, gene mutation). The age of onset varies greatly, from infant to adulthood. In addition, the slow and variable rates of disease progression in patients with HSP represent a substantial challenge for informative assessment of therapeutic efficacy. To address this, we are undertaking a prospective cohort study to investigate genetic–clinical characteristics, find surrogates for monitoring disease progress and identify clinical readouts for treatment.Methods and analysisIn this case-control cohort study, we will enrol 200 patients with HSP and 200 healthy individuals in parallel. Participants will be continuously assessed for 3 years at 12-month intervals. Six aspects, including clinical signs, genetic spectrum, cognitive competence, MRI features, potential biochemical indicators and nerve electrophysiological factors, will be assessed in detail. This study will observe clinical manifestations and disease severity based on different molecular mechanisms, including oxidative stress, cholesterol metabolism and microtubule dynamics, all of which have been proposed as potential treatment targets or modalities. The analysis will also assess disease progression in different types of HSPs and cellular pathways with a longitudinal study using t tests and χ2 tests.Ethics and disseminationThe study was granted ethics committee approval by the first affiliated hospital of Fujian Medical University (MRCTA, ECFAH of FMU (2019)194) in 2019. Findings will be disseminated via presentations and peer-reviewed publications. Dissemination will target different audiences, including national stakeholders, researchers from different disciplines and the general public.Trial registration numberNCT04006418.
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