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While whole genome sequencing and long-read sequencing have become widely available, more and more focuses are on noncoding expanded repeats. Indeed, more than half of noncoding repeat expansions related to diseases have been identified in the five years. An exciting aspect of the progress in this field is an identification of a phenomenon called repeat motif–phenotype correlation. Repeat motif–phenotype correlation in noncoding repeat expansion diseases is first found in benign adult familial myoclonus epilepsy. The concept is extended in the research of CGG repeat expansion diseases. In this review, we focus on newly identified CGG repeat expansion diseases, update the concept of repeat motif–phenotype correlation in CGG repeat expansion diseases, and propose a clinical concept of FNOP (fragile X-associated tremor/ataxia syndrome, neuronal intranuclear inclusion disease, and oculopharyngodistal myopathy)-spectrum disorder, which shares clinical features and thus probably share some common disease pathophysiology, to further facilitate discussion and progress in this field.

Some hereditary transthyretin (ATTRv) amyloidosis patients are misdiagnosed as Charcot-Marie-Tooth disease (CMT) at onset. We assess the findings to identify ATTRv amyloidosis among patients with suspected CMT to screen transthyretin gene variants for treatments.We assessed clinical, cerebrospinal fluid, and electrophysiological findings by comparing ATTRv amyloidosis patients with suspected CMT (n = 10) and CMT patients (n = 489).The median (interquartile range) age at onset of neurological symptoms was 69 (64.2-70) years in the ATTRv amyloidosis vs 12 (5-37.2) years in CMT group (Mann-Whitney U, p 0.01). The proportion of patients with initial sensory symptoms was 70% in the ATTRv amyloidosis group vs 7.1% in CMT group (Fisher's exact, p 0.01). The proportion of patients with histories of suspected chronic inflammatory demyelinating polyneuropathy (CIDP) were 50% in the ATTRv amyloidosis group vs 8.7% in CMT group (Fisher's exact, p .01). Other measures and outcomes were not different between the two groups. Five of the six patients with ATTRv amyloidosis received treatment and survived.For effective treatments, the transthyretin gene should be screened in patients with suspected CMT with old age at onset of neurological symptoms, initial sensory symptoms, and histories of suspected CIDP.

Hereditary spastic paraplegias (HSPs) comprise a group of neurodegenerative disorders characterized by weakness and leg spasticity. LYST is responsible for Chediak-Higashi syndrome (CHS), which exhibits partial oculocutaneous albinism, primary immunodeficiency, and bleeding tendency in childhood. Although neurological symptoms of CHS also appear in adulthood, a phenotype of spastic paraplegia has rarely been reported in CHS. In this study, we investigated LYST mutations in 387 HSP patients through the Japan Spastic Paraplegia Research Consortium to clarify the frequency of LYST mutations in HSP, finding six adult patients with LYST mutations in four HSP families. They exhibited intellectual disability, cerebellar ataxia, neuropathy, and pyramidal signs. Meanwhile, only 15 patients with CHS in childhood have been revealed in a decade by a nationwide survey in Japan. Thus, LYST mutations might indicate a HSP phenotype in a considerable number of adult patients with CHS.

Loss-of-function (LoF) variants in NEK1 have recently been reported to be associated with amyotrophic lateral sclerosis (ALS). In this study, we investigated the association of NEK1 LoF variants with an increased risk of sporadic ALS (SALS) and the clinical characteristics of patients with SALS carrying LoF variants in a Japanese case series. Whole-exome sequencing analysis was performed for a series of 446 SALS patients in whom pathogenic variants in familial ALS-causative genes have not been identified and 1163 healthy control subjects in our Japanese series. We evaluated LoF variants, defined as nonsense, splice-site disrupting single-nucleotide variants (SNVs), or short insertion/deletion (indel) variants predicted to cause frameshifts in NEK1. We identified seven NEK1 LoF variants in patients with SALS (1.57%), whereas only one was identified in control subjects (0.086%) (P = 0.00073, Fisher's exact test). This finding is consistent with those in recent reports from other regions in the world. In conclusion, we demonstrated that NEK1 LoF variants are also associated with an increased risk of SALS in the Japanese population.

Mutations in neurofilament genes have been linked to several neuromuscular disorders. The neurofilament heavy (NEFH) gene was identified as the causative gene of Charcot-Marie-Tooth disease type 2CC (CMT2CC) in 2016, with a toxic gain of function mechanism caused by the translation and aggregation of cryptic amyloidogenic element (CAE) in the 3' untranslated region (UTR). But the NEFH-related clinical and genetic spectrums are still unclear in Japan.We analyzed all variants in the NEFH gene from our in-house whole-exome sequencing data, established from Japanese nationwide patients with neuromuscular disorders, including Charcot-Marie-Tooth (CMT) disease and spinal muscular atrophy (SMA).We identified a c.3017dup (p.Pro1007Alafs*56) variant in NEFH from three families clinically diagnosed with CMT, and one family with SMA. In addition to the patients presented with typical peripheral neuropathies, pyramidal signs were observed from one CMT patient. Whereas the SMA patients showed severe characteristic weakness of triceps brachii and quadriceps femoris. All of these four families reside in Kagoshima Prefecture of Japan, and a following haplotype analysis strongly suggests a founder effect.This is the original report referring to a founder mutation in NEFH. The clinical diversity in our study, comprising CMT, with or without pyramidal signs, and SMA, suggest an extensive involvement of peripheral nerve, anterior horn cells, or both. Our findings broaden the phenotypic spectrum of NEFH-related disorders.

SPG31 is an autosomal dominant hereditary spastic paraplegia caused by pathogenic variants in the receptor expression-enhancing protein 1 (REEP1) gene. We analyzed 488 DNA samples from unrelated HSP patients collected by Japan Spastic Paraplegia Research Consortium and found 15 Japanese SPG31 families. We investigated each family and found a total of 25 individuals with REEP1 variants (comprising 22 patients and three asymptomatic carriers). Fourteen REEP1 variants (five missense, three nonsense, four frameshift, one splice site, and one large deletion) including 11 novel ones were detected. Seventy percent of the patients (14 of 20) showed a pure form and the others (6 of 20) showed a complicated form with peripheral neuropathy. Fifty percent of the patients had neurological symptoms before the age of 10 and 20% of them at age 41-50. The mean age of onset was 19.6 ± 18.7 (from 5 to 67, n = 15) years for males and 32.8 ± 24.7 (from 4 to 60, n = 5) years for females. Although the difference was not statistically significant (p = 0.38, Mann-Whitney U test), males tended to have an earlier age of onset. Moreover, all three asymptomatic carriers were female. We investigated additional factors as to phenotypic appearance in one family with apparent intrafamilial variability in age at onset and clinical severity, but no additional factors including gene variants could be found. This is the first report of clinical and genetic findings of SPG31 in Japan, which may lead to further studies of the genotype-phenotype correlation of SPG31.

Hereditary spastic paraplegias (HSPs) comprise a group of neurodegenerative disorders characterized by weakness and leg spasticity. LYST is responsible for Chédiak-Higashi syndrome (CHS), which exhibits partial oculocutaneous albinism, primary immunodeficiency, and bleeding tendency in childhood. Although neurological symptoms of CHS also appear in adulthood, a phenotype of spastic paraplegia has rarely been reported in CHS. In this study, we investigated LYST mutations in 387 HSP patients through the Japan Spastic Paraplegia Research Consortium to clarify the frequency of LYST mutations in HSP, finding six adult patients with LYST mutations in four HSP families. They exhibited intellectual disability, cerebellar ataxia, neuropathy, and pyramidal signs. Meanwhile, only 15 patients with CHS in childhood have been revealed in a decade by a nationwide survey in Japan. Thus, LYST mutations might indicate a HSP phenotype in a considerable number of adult patients with CHS.

Adrenoleukodystrophy (ALD) is an X-linked disease that affects primarily the white matter of the central nervous system and adrenal cortex. A correlation between genotypes and phenotypes has not been observed. Here, we present two Japanese siblings with a novel missense variant (c.1887T > G) in the ABCD1 gene who presented with different clinical phenotypes, i.e., adolescent cerebral and cerebello-brainstem types. We also review the literature focusing on the variation in the clinical phenotypes within ALD families. In our review, 61.9% of sibling pairs presented with the same clinical type of ALD and 59.1% of sibling pairs presented with a similar age of onset. Conversely, 15.4% of sibling pairs had a similar age of onset, but different clinical types of ALD. To observe the correlation between genotypes and phenotypes, it is important to diagnose early and to accumulate reports describing age of onset, first onset symptom, and progression of the symptom.

Adrenoleukodystrophy (ALD) is an X-linked disease that affects primarily the white matter of the central nervous system and adrenal cortex. A correlation between genotypes and phenotypes has not been observed. Here, we present two Japanese siblings with a novel missense variant (c.1887T G) in the ABCD1 gene who presented with different clinical phenotypes, i.e., adolescent cerebral and cerebello-brainstem types. We also review the literature focusing on the variation in the clinical phenotypes within ALD families. In our review, 61.9% of sibling pairs presented with the same clinical type of ALD and 59.1% of sibling pairs presented with a similar age of onset. Conversely, 15.4% of sibling pairs had a similar age of onset, but different clinical types of ALD. To observe the correlation between genotypes and phenotypes, it is important to diagnose early and to accumulate reports describing age of onset, first onset symptom, and progression of the symptom.

The tropomyosin-receptor kinase fused gene (TFG) has recently been implicated in several distinct hereditary disorders, including the autosomal-recessive form of complicated hereditary spastic paraplegia called SPG57. Previously, three homozygous variants of the TFG gene were reported in five families with SPG57, in which early onset spastic paraplegia, optic atrophy, and peripheral neuropathy were variably identified. Here, we present the first Japanese patient with SPG57, and have added a homozygous p.Ile66Thr variant as the fourth SPG57 genotype.

Hereditary spastic paraplegias (HSPs) are characterized by various inherited disorders in which weakness and spasticity of the lower extremities are the predominant symptoms. Recently, HSP caused by ALDH18A1 mutations has been reported as SPG9 with autosomal dominant (SPG9A) and autosomal recessive (SPG9B) transmission. In this study, we obtained clinical and genetic findings in two Japanese families with SPG9B. One family had a novel compound heterozygous mutation (c.1321 C > T/c.1994G > A) in the ALDH18A1 gene. The other family had a homozygous mutation (c.383 G > A/c.383 G > A) in the ALDH18A1 gene. To date, only two SPG9B families with ALDH18A1 mutations have been reported. This is the first report of SPG9 in non-Caucasians. Furthermore, we found cerebellar ataxia in one family, although cerebellar ataxia has not been reported in SPG9B so far. SPG9B might involve a complicated HSP including cerebellar ataxia and cognitive impairment. This study expands the clinical and genetic spectrum of ALDH18A1-related disorders.

MODY 5 and 6 have been shown to be low-penetrant MODYs. As the genetic background of unknown MODY is assumed to be similar, a new analytical strategy is applied here to elucidate genetic predispositions to unknown MODY. We examined to find whether there are major MODY gene loci remaining to be identified using SNP linkage analysis in Japanese. Whole-exome sequencing was performed with seven families with typical MODY. Candidates for novel MODY genes were examined combined with in silico network analysis. Some peaks were found only in either parametric or non-parametric analysis; however, none of these peaks showed a LOD score greater than 3.7, which is approved to be the significance threshold of evidence for linkage. Exome sequencing revealed that three mutated genes were common among 3 families and 42 mutated genes were common in two families. Only one of these genes, MYO5A, having rare amino acid mutations p.R849Q and p.V1601G, was involved in the biological network of known MODY genes through the intermediary of the INS. Although only one promising candidate gene, MYO5A, was identified, no novel, high penetrant MODY genes might remain to be found in Japanese MODY.

Spinal muscular atrophy (SMA) is an autosomal recessive neurodegenerative disorder caused by survival motor neuron gene mutations. Variant forms of SMA accompanied by additional clinical presentations have been classified as atypical SMA and are thought to be caused by variants in as yet unidentified causative genes. Here, we presented the clinical findings of two siblings with an SMA variant followed by progressive cerebral atrophy, and the results of whole-exome sequencing analyses of the family quartet that was performed to identify potential causative variants. We identified two candidate homozygous missense variants, R942Q in the tubulin-folding cofactor D (TBCD) gene and H250Q in the bromo-adjacent homology domain and coiled-coil containing 1 (BAHCC1) gene, located on chromosome 17q25.3 with an interval of 1.4 Mbp. The in silico analysis of both variants suggested that TBCD rather than BAHCC1 was likely the pathogenic gene (TBCD sensitivity, 0.68; specificity, 0.97; BAHCC1 sensitivity, 1.00; specificity, 0.00). Thus, our results show that TBCD is a likely novel candidate gene for atypical SMA with progressive cerebral atrophy. TBCD is predicted to have important functions on tubulin integrity in motor neurons as well as in the central nervous system.

The originally published version of this article contained an error in Fig. 1 and Table 2. The correct figure and table of this article should have read as below. This has now been corrected in the PDF and HTML versions of the article. The authors apologize for any inconvenience caused.

We aimed to find a new causative gene and elucidate the molecular mechanisms underlying a new type of hereditary spastic paraplegia (HSP). Patients with HSP were recruited from the Japan Spastic Paraplegia Research Consortium (JASPAC). Exome sequencing of genomic DNA from patients in four families was carried out, followed by Sanger sequencing of the UBAP1 gene. A mouse homolog of one UBAP1 frameshift mutation carried by one of the patients was created as a disease model. Functional properties of the UBAP1 wild type and UBAP1-mutant in mouse hippocampus neurons were examined. We identified three novel heterozygous loss of function mutations (c.425_426delAG, c.312delC, and c.535G>T) in the UBAP1 gene as the genetic cause of a new type of HSP (SPG80). All the patients presented identical clinical features of a pure type of juvenile-onset HSP. Functional studies on mouse hippocampal neurons revealed that the C-terminal deletion UBAP1-mutant of our disease model had lost its ability to bind ubiquitin in vitro. Overexpression of the UBAP1 wild type interacts directly with ubiquitin on enlarged endosomes, while the UBAP1-mutant cannot be recruited to endosome membranes. Our study demonstrated that mutations in the UBAP1 gene cause a new type of HSP and elucidated its pathogenesis. The full-length UBAP1 protein is involved in endosomal dynamics in neurons, while loss of UBAP1 function may perturb endosomal fusion and sorting of ubiquitinated cargos. These effects could be more prominent in neurons, thereby giving rise to the phenotype of a neurodegenerative disease such as HSP.

Whole-genome and -exome resequencing using next-generation sequencers is a powerful approach for identifying genomic variations that are associated with diseases. However, systematic strategies for prioritizing causative variants from many candidates to explain the disease phenotype are still far from being established, because the population-specific frequency spectrum of genetic variation has not been characterized. Here, we have collected exomic genetic variation from 1208 Japanese individuals through a collaborative effort, and aggregated the data into a prevailing catalog. In total, we identified 156 622 previously unreported variants. The allele frequencies for the majority (88.8%) were lower than 0.5% in allele frequency and predicted to be functionally deleterious. In addition, we have constructed a Japanese-specific major allele reference genome by which the number of unique mapping of the short reads in our data has increased 0.045% on average. Our results illustrate the importance of constructing an ethnicity-specific reference genome for identifying rare variants. All the collected data were centralized to a newly developed database to serve as useful resources for exploring pathogenic variations. Public access to the database is available at http://www.genome.med.kyoto-u.ac.jp/SnpDB/.

PLA2G6-associated neurodegeneration (PLAN) comprises heterogeneous neurodegenerative disorders, including infantile neuroaxonal dystrophy, neurodegeneration with brain iron accumulation 2B, and Parkinson disease 14 (PARK14). In addition, very recently, PLA2G6 mutations have been reported to represent a phenotype of hereditary spastic paraplegia (HSP). In this study, we screened 383 HSP families to clarify the frequency of PLA2G6 mutations in the Japan Spastic Paraplegia Research Consortium, and revealed the clinical characteristics of HSP with PLA2G6 mutations. We found three families with compound heterozygous mutations of the PLA2G6 gene, c.517 C T/c.1634A G, c.662 T C/c.991 G T, and c.1187-2 A G/c.1933C T, and one family with a homozygous mutation of the PLA2G6 gene, c.1904G A/c.1904G A. All three families with compound heterozygous mutations presented a uniform phenotype of a complicated form of HSP with infantile/child-onset spastic paraplegia, cerebellar ataxia, and mental retardation. On the other hand, the family with a homozygous mutation presented a late-onset complicated form of HSP with parkinsonism. This study may extend the clinical and genetic findings for PLAN.

On whole-exome sequencing, a novel compound heterozygous mutation (c.2923A>G/c.3523_3524insTGTCCG, p.T975A/p.1175_1176insVS) and a novel homozygous one (c.3534G>C, p.W1178C) in the PNPLA6 gene were identified in sporadic and familial Japanese patients with Boucher-Neuhauser syndrome (BNS), respectively. However, we did not find any mutations in the PNPLA6 gene in 88 patients with autosomal recessive hereditary spastic paraplegia (ARHSP). Our study confirmed the earlier report that a PNPLA6 mutation causes BNS. This is the first report on PNPLA6 mutations in non-Caucasian patients. Meanwhile, PNPLA6 mutations might be extremely rare in Japanese ARHSP patients. Moreover, we first found hypersegmented neutrophils in two BNS patients with PNPLA6 mutations.

Hereditary spastic paraplegia (HSP) is one of the most genetically heterogeneous neurodegenerative disorders characterized by progressive spasticity and pyramidal weakness of lower limbs. Because >30 causative genes have been identified, screening of multiple genes is required for establishing molecular diagnosis of individual patients with HSP. To elucidate molecular epidemiology of HSP in the Japanese population, we have conducted mutational analyses of 16 causative genes of HSP (L1CAM, PLP1, ATL1, SPAST, CYP7B1, NIPA1, SPG7, KIAA0196, KIF5A, HSPD1, BSCL2, SPG11, SPG20, SPG21, REEP1 and ZFYVE27) using resequencing microarrays, array-based comparative genomic hybridization and Sanger sequencing. The mutational analysis of 129 Japanese patients revealed 49 mutations in 46 patients, 32 of which were novel. Molecular diagnosis was accomplished for 67.3% (33/49) of autosomal dominant HSP patients. Even among sporadic HSP patients, mutations were identified in 11.1% (7/63) of them. The present study elucidated the molecular epidemiology of HSP in the Japanese population and further broadened the mutational and clinical spectra of HSP.

We have recently found that multiple rare variants of the glucocerebrosidase gene (GBA) confer a robust risk for Parkinson disease, supporting the 'common disease-multiple rare variants' hypothesis. To develop an efficient method of identifying rare variants in a large number of samples, we applied multiplexed resequencing using a next-generation sequencer to identification of rare variants of GBA. Sixteen sets of pooled DNAs from six pooled DNA samples were prepared. Each set of pooled DNAs was subjected to polymerase chain reaction to amplify the target gene (GBA) covering 6.5 kb, pooled into one tube with barcode indexing, and then subjected to extensive sequence analysis using the SOLiD System. Individual samples were also subjected to direct nucleotide sequence analysis. With the optimization of data processing, we were able to extract all the variants from 96 samples with acceptable rates of false-positive single-nucleotide variants.