Satoko Miyatake

Mutations affecting components of the SWI/SNF complex cause Coffin-Siris syndrome.

By exome sequencing, we found de novo SMARCB1 mutations in two of five individuals with typical Coffin-Siris syndrome (CSS), a rare autosomal dominant anomaly syndrome. As SMARCB1 encodes a subunit of the SWItch/Sucrose NonFermenting (SWI/SNF) complex, we screened 15 other genes encoding subunits of this complex in 23 individuals with CSS. Twenty affected individuals (87%) each had a germline mutation in one of six SWI/SNF subunit genes, including SMARCB1, SMARCA4, SMARCA2, SMARCE1, ARID1A and ARID1B.

Homozygous c.14576G>A variant of RNF213 predicts early-onset and severe form of moyamoya disease

Objective: RNF213 was recently reported as a susceptibility gene for moyamoya disease (MMD). Our aim was to clarify the correlation between the RNF213 genotype and MMD phenotype. Methods: The entire coding region of the RNF213 gene was sequenced in 204 patients with MMD, and corresponding variants were checked in 62 pairs of parents, 13 mothers and 4 fathers of the patients, and 283 normal controls. Clinical information was collected. Genotype-phenotype correlations were statistically analyzed. Results: The c.14576G>A variant was identified in 95.1% of patients with familial MMD, 79.2% of patients with sporadic MMD, and 1.8% of controls, thus confirming its association with MMD, with an odds ratio of 259 and p < 0.001 for either heterozygotes or homozygotes. Homozygous c.14576G>A was observed in 15 patients but not in the controls and unaffected parents. The incidence rate for homozygotes was calculated to be >78%. Homozygotes had a significantly earlier age at onset compared with heterozygotes or wild types (median age at onset 3, 7, and 8 years, respectively). Of homozygotes, 60% were diagnosed with MMD before age 4, and all had infarctions as the first symptom. Infarctions at initial presentation and involvement of posterior cerebral arteries, both known as poor prognostic factors for MMD, were of significantly higher frequency in homozygotes than in heterozygotes and wild types. Variants other than c.14576G>A were not associated with clinical phenotypes. Conclusions: The homozygous c.14576G>A variant in RNF213 could be a good DNA biomarker for predicting the severe type of MMD, for which early medical/surgical intervention is recommended, and may provide a better monitoring and prevention strategy.

Mutations in KLHL40 Are a Frequent Cause of Severe Autosomal-Recessive Nemaline Myopathy

Nemaline myopathy (NEM) is a common congenital myopathy. At the very severe end of the NEM clinical spectrum are genetically unresolved cases of autosomal-recessive fetal akinesia sequence. We studied a multinational cohort of 143 severe-NEM-affected families lacking genetic diagnosis. We performed whole-exome sequencing of six families and targeted gene sequencing of additional families. We identified 19 mutations in KLHL40 (kelch-like family member 40) in 28 apparently unrelated NEM kindreds of various ethnicities. Accounting for up to 28% of the tested individuals in the Japanese cohort, KLHL40 mutations were found to be the most common cause of this severe form of NEM. Clinical features of affected individuals were severe and distinctive and included fetal akinesia or hypokinesia and contractures, fractures, respiratory failure, and swallowing difficulties at birth. Molecular modeling suggested that the missense substitutions would destabilize the protein. Protein studies showed that KLHL40 is a striated-muscle-specific protein that is absent in KLHL40-associated NEM skeletal muscle. In zebrafish, klhl40a and klhl40b expression is largely confined to the myotome and skeletal muscle, and knockdown of these isoforms results in disruption of muscle structure and loss of movement. We identified KLHL40 mutations as a frequent cause of severe autosomal-recessive NEM and showed that it plays a key role in muscle development and function. Screening of KLHL40 should be a priority in individuals who are affected by autosomal-recessive NEM and who present with prenatal symptoms and/or contractures and in all Japanese individuals with severe NEM.

Phenotypic Spectrum of COL4A1 Mutations: Porencephaly to Schizencephaly

Recently, COL4A1 mutations have been reported in porencephaly and other cerebral vascular diseases, often associated with ocular, renal, and muscular features. In this study, we aimed to clarify the phenotypic spectrum and incidence of COL4A1 mutations.

Dominant mutations in ORAI1 cause tubular aggregate myopathy with hypocalcemia via constitutive activation of store-operated Ca2+ channels

The store-operated Ca(2+) release-activated Ca(2+) (CRAC) channel is activated by diminished luminal Ca(2+) levels in the endoplasmic reticulum and sarcoplasmic reticulum (SR), and constitutes one of the major Ca(2+) entry pathways in various tissues. Tubular aggregates (TAs) are abnormal structures in the skeletal muscle, and although their mechanism of formation has not been clarified, altered Ca(2+) homeostasis related to a disordered SR is suggested to be one of the main contributing factors. TA myopathy is a hereditary muscle disorder that is pathologically characterized by the presence of TAs. Recently, dominant mutations in the STIM1 gene, encoding a Ca(2+) sensor that controls CRAC channels, have been identified to cause tubular aggregate myopathy (TAM). Here, we identified heterozygous missense mutations in the ORAI1 gene, encoding the CRAC channel itself, in three families affected by dominantly inherited TAM with hypocalcemia. Skeletal myotubes from an affected individual and HEK293 cells expressing mutated ORAI1 proteins displayed spontaneous extracellular Ca(2+) entry into cells without diminishment of luminal Ca(2+) or the association with STIM1. Our results indicate that STIM1-independent activation of CRAC channels induced by dominant mutations in ORAI1 cause altered Ca(2+) homeostasis, resulting in TAM with hypocalcemia.

SMOC1 is essential for ocular and limb development in humans and mice.

Microphthalmia with limb anomalies (MLA) is a rare autosomal-recessive disorder, presenting with anophthalmia or microphthalmia and hand and/or foot malformation. We mapped the MLA locus to 14q24 and successfully identified three homozygous (one nonsense and two splice site) mutations in the SPARC (secreted protein acidic and rich in cysteine)-related modular calcium binding 1 (SMOC1) in three families. Smoc1 is expressed in the developing optic stalk, ventral optic cup, and limbs of mouse embryos. Smoc1 null mice recapitulated MLA phenotypes, including aplasia or hypoplasia of optic nerves, hypoplastic fibula and bowed tibia, and syndactyly in limbs. A thinned and irregular ganglion cell layer and atrophy of the anteroventral part of the retina were also observed. Soft tissue syndactyly, resulting from inhibited apoptosis, was related to disturbed expression of genes involved in BMP signaling in the interdigital mesenchyme. Our findings indicate that SMOC1/Smoc1 is essential for ocular and limb development in both humans and mice.

De novo SOX11 mutations cause Coffin–Siris syndrome

Coffin-Siris syndrome (CSS) is a congenital disorder characterized by growth deficiency, intellectual disability, microcephaly, characteristic facial features and hypoplastic nails of the fifth fingers and/or toes. We previously identified mutations in five genes encoding subunits of the BAF complex, in 55% of CSS patients. Here we perform whole-exome sequencing in additional CSS patients, identifying de novo SOX11 mutations in two patients with a mild CSS phenotype. sox11a/b knockdown in zebrafish causes brain abnormalities, potentially explaining the brain phenotype of CSS. SOX11 is the downstream transcriptional factor of the PAX6-BAF complex, highlighting the importance of the BAF complex and SOX11 transcriptional network in brain development.

Clinical correlations of mutations affecting six components of the SWI/SNF complex: Detailed description of 21 patients and a review of the literature

Mutations in the components of the SWItch/sucrose nonfermentable (SWI/SNF)‐like chromatin remodeling complex have recently been reported to cause Coffin–Siris syndrome (CSS), Nicolaides–Baraitser syndrome (NCBRS), and ARID1B‐related intellectual disability (ID) syndrome. We detail here the genotype‐phenotype correlations for 85 previously published and one additional patient with mutations in the SWI/SNF complex: four with SMARCB1 mutations, seven with SMARCA4 mutations, 37 with SMARCA2 mutations, one with an SMARCE1 mutation, three with ARID1A mutations, and 33 with ARID1B mutations. The mutations were associated with syndromic ID and speech impairment (severe/profound in SMARCB1, SMARCE1, and ARID1A mutations; variable in SMARCA4, SMARCA2, and ARID1B mutations), which was frequently accompanied by agenesis or hypoplasia of the corpus callosum. SMARCB1 mutations caused “classical” CSS with typical facial “coarseness” and significant digital/nail hypoplasia. SMARCA4 mutations caused CSS without typical facial coarseness and with significant digital/nail hypoplasia. SMARCA2 mutations caused NCBRS, typically with short stature, sparse hair, a thin vermillion of the upper lip, an everted lower lip and prominent finger joints. A SMARCE1 mutation caused CSS without typical facial coarseness and with significant digital/nail hypoplasia. ARID1A mutations caused the most severe CSS with severe physical complications. ARID1B mutations caused CSS without typical facial coarseness and with mild digital/nail hypoplasia, or caused syndromic ID. Because of the common underlying mechanism and overlapping clinical features, we propose that these conditions be referred to collectively as “SWI/SNF‐related ID syndromes”.

Pathogenic Variants in PIGG Cause Intellectual Disability with Seizures and Hypotonia

Glycosylphosphatidylinositol (GPI) is a glycolipid that anchors >150 various proteins to the cell surface. At least 27 genes are involved in biosynthesis and transport of GPI-anchored proteins (GPI-APs). To date, mutations in 13 of these genes are known to cause inherited GPI deficiencies (IGDs), and all are inherited as recessive traits. IGDs mainly manifest as intellectual disability, epilepsy, coarse facial features, and multiple organ anomalies. These symptoms are caused by the decreased surface expression of GPI-APs or by structural abnormalities of GPI. Here, we present five affected individuals (from two consanguineous families from Egypt and Pakistan and one non-consanguineous family from Japan) who show intellectual disability, hypotonia, and early-onset seizures. We identified pathogenic variants in PIGG, a gene in the GPI pathway. In the consanguineous families, homozygous variants c.928C>T (p.Gln310(∗)) and c.2261+1G>C were found, whereas the Japanese individual was compound heterozygous for c.2005C>T (p.Arg669Cys) and a 2.4 Mb deletion involving PIGG. PIGG is the enzyme that modifies the second mannose with ethanolamine phosphate, which is removed soon after GPI is attached to the protein. Physiological significance of this transient modification has been unclear. Using B lymphoblasts from affected individuals of the Egyptian and Japanese families, we revealed that PIGG activity was almost completely abolished; however, the GPI-APs had normal surface levels and normal structure, indicating that the pathogenesis of PIGG deficiency is not yet fully understood. The discovery of pathogenic variants in PIGG expands the spectrum of IGDs and further enhances our understanding of this etiopathogenic class of intellectual disability.

Expanding the phenotypic spectrum of TUBB4A-associated hypomyelinating leukoencephalopathies

Objective: We performed whole-exome sequencing analysis of patients with genetically unsolved hypomyelinating leukoencephalopathies, identifying 8 patients with TUBB4A mutations and allowing the phenotypic spectrum of TUBB4A mutations to be investigated. Methods: Fourteen patients with hypomyelinating leukoencephalopathies, 7 clinically diagnosed with hypomyelination with atrophy of the basal ganglia and cerebellum (H-ABC), and 7 with unclassified hypomyelinating leukoencephalopathy, were analyzed by whole-exome sequencing. The effect of the mutations on microtubule assembly was examined by mapping altered amino acids onto 3-dimensional models of the αβ-tubulin heterodimer. Results: Six heterozygous missense mutations in TUBB4A, 5 of which are novel, were identified in 8 patients (6/7 patients with H-ABC [the remaining patient is an atypical case] and 2/7 patients with unclassified hypomyelinating leukoencephalopathy). In 4 cases with parental samples available, the mutations occurred de novo. Analysis of 3-dimensional models revealed that the p.Glu410Lys mutation, identified in patients with unclassified hypomyelinating leukoencephalopathy, directly impairs motor protein and/or microtubule-associated protein interactions with microtubules, whereas the other mutations affect longitudinal interactions for maintaining αβ-tubulin structure, suggesting different mechanisms in tubulin function impairment. In patients with the p.Glu410Lys mutation, basal ganglia atrophy was unobserved or minimal although extrapyramidal features were detected, suggesting its functional impairment. Conclusions: TUBB4A mutations cause typical H-ABC. Furthermore, TUBB4A mutations associate cases of unclassified hypomyelinating leukoencephalopathies with morphologically retained but functionally impaired basal ganglia, suggesting that TUBB4A-related hypomyelinating leukoencephalopathies encompass a broader clinical spectrum than previously expected. Extrapyramidal findings may be a key for consideration of TUBB4A mutations in hypomyelinating leukoencephalopathies.