Hemizygous variants in protein phosphatase 1 regulatory subunit 3F (PPP1R3F) are associated with a neurodevelopmental disorder characterized by developmental delay, intellectual disability and autistic features.

Protein phosphatase 1 regulatory subunit 3F (PPP1R3F) is a member of the glycogen targeting subunits (GTSs), which belong to the large group of regulatory subunits of protein phosphatase 1 (PP1), a major eukaryotic serine/threonine protein phosphatase that regulates diverse cellular processes. Here, we describe the identification of hemizygous variants in PPP1R3F associated with a novel X-linked recessive neurodevelopmental disorder in 13 unrelated individuals. This disorder is characterized by developmental delay, mild intellectual disability, neurobehavioral issues such as autism spectrum disorder, seizures and other neurological findings including tone, gait and cerebellar abnormalities. PPP1R3F variants segregated with disease in affected hemizygous males that inherited the variants from their heterozygous carrier mothers. We show that PPP1R3F is predominantly expressed in brain astrocytes and localizes to the endoplasmic reticulum in cells. Glycogen content in PPP1R3F knockout astrocytoma cells appears to be more sensitive to fluxes in extracellular glucose levels than in wild-type cells, suggesting that PPP1R3F functions in maintaining steady brain glycogen levels under changing glucose conditions. We performed functional studies on nine of the identified variants and observed defects in PP1 binding, protein stability, subcellular localization and regulation of glycogen metabolism in most of them. Collectively, the genetic and molecular data indicate that deleterious variants in PPP1R3F are associated with a new X-linked disorder of glycogen metabolism, highlighting the critical role of GTSs in neurological development. This research expands our understanding of neurodevelopmental disorders and the role of PP1 in brain development and proper function.

Genetic variations in NLRP3 and NLRP12 genes in adult-onset patients with autoinflammatory diseases: a comparative study.

Objectives: Cryopyrin-associated periodic syndrome or NLRP3-associated autoinflammatory disease (NLRP3-AID) and NLRP12-AID are both Mendelian disorders with autosomal dominant inheritance. Both diseases are rare, primarily reported in the pediatric population, and are thought to be phenotypically indistinguishable. We provide the largest cohort of adult-onset patients and compared these diseases and the gene variant frequency to population controls. Methods: A cohort of adult patients with AIDs were retrospectively studied. All underwent molecular testing for periodic fever syndrome gene panels after extensive and negative workups for systemic autoimmune and other related diseases. Patients were divided into Group 1- NLRP3-AID patients with NLRP3 variants (N=15), Group 2- NLRP12-AID with NLRP12 variants (N=14) and Group 3- both NLRP3 and NLRP12 (N=9) variants. Exome sequence data of two large control populations including the ARIC study were used to compare gene variant distribution and frequency. Results: All 38 patients were Caucasian with women accounting for 82%. Median age at diagnosis was 41 ± 23 years and the disease duration at diagnosis was 14 ± 13 years. We identified statistically significant differences between the groups, notably that gastrointestinal symptoms as well as evaluations for same were significantly more frequent in patients with NLRP12 variants, and headaches/dizziness were less common among the NLRP12 patients. Livedo reticularis was noted in four patients, exclusively among NLRP12 carriers. Over 50% of patients in Groups 1 and 2 carry low-frequency disease-associated variants, while the remaining carry rare variants. We unprecedently identified digenic variants, i.e., the coexistence of NLRP3 and NLRP12, which were either both low frequency or low frequency/rare. Allele frequencies of all variants identified in our cohort were either absent or significantly lower in the control populations, further strengthening the evidence of susceptibility of these variants to SAID phenotypes. Conclusion: Our comparative study shows that both NLRP3-AID and NLRP12-AID share similar clinical phenotypes, yet there are significant differences between them with regard to gastrointestinal and neurological symptoms. A spectrum of high to low genetic variations in both genes can contribute to SAID individually or in combination.

Bi-allelic variants in neuronal cell adhesion molecule cause a neurodevelopmental disorder characterized by developmental delay, hypotonia, neuropathy/spasticity.

Cell adhesion molecules are membrane-bound proteins predominantly expressed in the central nervous system along principal axonal pathways with key roles in nervous system development, neural cell differentiation and migration, axonal growth and guidance, myelination, and synapse formation. Here, we describe ten affected individuals with bi-allelic variants in the neuronal cell adhesion molecule NRCAM that lead to a neurodevelopmental syndrome of varying severity; the individuals are from eight families. This syndrome is characterized by developmental delay/intellectual disability, hypotonia, peripheral neuropathy, and/or spasticity. Computational analyses of NRCAM variants, many of which cluster in the third fibronectin type III (Fn-III) domain, strongly suggest a deleterious effect on NRCAM structure and function, including possible disruption of its interactions with other proteins. These findings are corroborated by previous in vitro studies of murine Nrcam-deficient cells, revealing abnormal neurite outgrowth, synaptogenesis, and formation of nodes of Ranvier on myelinated axons. Our studies on zebrafish nrcamaΔ mutants lacking the third Fn-III domain revealed that mutant larvae displayed significantly altered swimming behavior compared to wild-type larvae (p < 0.03). Moreover, nrcamaΔ mutants displayed a trend toward increased amounts of α-tubulin fibers in the dorsal telencephalon, demonstrating an alteration in white matter tracts and projections. Taken together, our study provides evidence that NRCAM disruption causes a variable form of a neurodevelopmental disorder and broadens the knowledge on the growing role of the cell adhesion molecule family in the nervous system.

From Disease Description and Gene Discovery to Functional Cell Pathway: A Decade-Long Journey for TMCO1.

A decade has passed since transmembrane coiled-coil domains 1 (TMCO1) defect syndrome was identified in 11 undiagnosed patients within the Old Order Amish of Northeastern Ohio-a disorder characterized by a distinctive craniofacial dysmorphism, skeletal anomalies and global developmental delay. Twenty seven patients, from diverse ethnic groups, have been reported with pathogenic TMCO1 variants now recognized to cause cerebrofaciothoracic dysplasia (CFTD). The implication of previously uncharacterized TMCO1 within disease has instigated a 10-year journey to understand the function of TMCO1 protein in Ca2+ homeostasis. TMCO1 is an ER Ca2+ leak channel which facilitates Ca2+ leak upon ER "overload" through the novel Ca2+ load activated Ca2+ mechanism. This mini-review brings together the clinical and scientific advances made since the discovery of TMCO1 deficiency in disease, including broadened phenotype, understanding of pathophysiology, and implications to patient management of TMCO1 defect syndrome.

Tissue-Biased Expansion of DNMT3A-Mutant Clones in a Mosaic Individual Is Associated with Conserved Epigenetic Erosion.

DNA methyltransferase 3A (DNMT3A) is the most commonly mutated gene in clonal hematopoiesis (CH). Somatic DNMT3A mutations arise in hematopoietic stem cells (HSCs) many years before malignancies develop, but difficulties in comparing their impact before malignancy with wild-type cells have limited the understanding of their contributions to transformation. To circumvent this limitation, we derived normal and DNMT3A mutant lymphoblastoid cell lines from a germline mosaic individual in whom these cells co-existed for nearly 6 decades. Mutant cells dominated the blood system, but not other tissues. Deep sequencing revealed similar mutational burdens and signatures in normal and mutant clones, while epigenetic profiling uncovered the focal erosion of DNA methylation at oncogenic regulatory regions in mutant clones. These regions overlapped with those sensitive to DNMT3A loss after DNMT3A ablation in HSCs and in leukemia samples. These results suggest that DNMT3A maintains a conserved DNA methylation pattern, the erosion of which provides a distinct competitive advantage to hematopoietic cells.

Dopa-Responsive Dystonia: A Male Patient Inherited a Novel GCH1 Deletion from an Asymptomatic Mother.

Dopa-responsive dystonia (DRD) is a complex genetic disorder with either autosomal dominant or autosomal recessive inheritance, with autosomal dominant being more frequent. Autosomal dominant DRD is known to be caused by mutations in the GCH1 gene, with incomplete penetrance frequently reported, particularly in males. Here, we report a male patient with DRD caused by exon 1 deletion in the GCH1 gene inherited from the asymptomatic mother. The patient had an atypical presentation, notably with no dystonia, and underwent extensive workup for a myriad of neuromuscular disorders before a low-dose L-dopa trial and confirmatory genetic testing were performed. Our experience with this family highlights an atypical presentation of DRD and prompts us to consider the genetic complexity of DRD.

Oral Ganglioside Supplement Improves Growth and Development in Patients with Ganglioside GM3 Synthase Deficiency.

Ganglioside GM3 synthase is a key enzyme involved in the biosynthesis of gangliosides. GM3 synthase deficiency (GM3D) causes an absence of GM3 and all downstream biosynthetic derivatives. The affected individuals manifest with severe irritability, intractable seizures, and profound intellectual disability. The current study is to assess the effects of an oral ganglioside supplement to patients with GM3D, particularly on their growth and development during early childhood. A total of 13 young children, 11 of them under 40 months old, received oral ganglioside supplement through a dairy product enriched in gangliosides, for an average of 34 months. Clinical improvements were observed in most children soon after the supplement was initiated. Significantly improved growth and development were documented in these subjects as average percentiles for weight, height, and occipitofrontal circumference increased in 1-2 months. Three children with initial microcephaly demonstrated significant catch-up head growth and became normocephalic. We also illustrated brief improvements in developmental and cognitive scores, particularly in communication and socialization domains through Vineland-II. However, all improvements seemed transient and gradually phased out after 12 months of supplementation. Gangliosides GM1 and GM3, although measureable in plasma during the study, were not significantly changed with ganglioside supplementation for up to 30 months. We speculate that the downstream metabolism of ganglioside biosynthesis is fairly active and the potential need for gangliosides in the human body is likely substantial. As we search for new effective therapies for GM3D, approaches to reestablish endogenous ganglioside supplies in the affected individuals should be considered.

Congenital Glucose-Galactose Malabsorption: A Case Report.

Congenital glucose-galactose malabsorption (CGGM) is a rare cause of intractable infantile diarrhea, with only a few hundred cases recognized worldwide. This life-threatening disorder must be considered in the differential diagnosis of an infant who presents with diarrhea and dehydration that fails to respond to standard therapy. The clinical and diagnostic course of an infant with recurrent episodes of watery diarrhea and hypernatremic dehydration found to be homozygous for a rare variant in the SLC5A1 gene, c.187C>T (p.R63X) is described.

Quantification of monosialogangliosides in human plasma through chemical derivatization for signal enhancement in LC-ESI-MS.

Gangliosides are found in abundance in the central nervous system of vertebrates. Their metabolic disruption and dysfunction are associated with various neurodegenerative disorders such as Alzheimer's disease and Parkinson's disease. In order to improve our understanding of the etiology of these diseases, analytical ganglioside assays with sufficient specificity and sensitivity in relevant biological matrices are required. In the present work we have developed and validated a reverse-phase ultra-performance liquid chromatography (UPLC)/tandem mass spectrometry (MS) method for determining monosialogangliosides GM1, GM2, and GM3 present in human plasma. Compared with our previous method, this method enhanced, by 15 fold, MS responses of the analytes by employing 2-(2-Pyridilamino)-ethylamine (PAEA) & 4-(4, 6-Dimethoxy-1, 3, 5-triazin-2-yl)-4-methylmorpholinium chloride (DMTMM)-based derivatization. The analytes and internal standards were derivatized with PAEA&DMTMM after extraction from plasma using a protein precipitation procedure. They were then purified using liquid-liquid partitioning. When the samples were then analyzed by UPLC-MS/MS with a multiple reaction monitoring (MRM) mode, we achieved superior sensitivity and specificity. This method was evaluated for extraction recovery, calibration linearity, precision, accuracy, and lower limit of quantification (LLOQ). The validated method was successfully applied to monitor monosialoganglioside levels in the plasma from patients with GM3 synthase deficiency. With significantly increased sensitivity, we have, for the first time, detected a significant amount of GM3 in the affected patients.

SAMHD1 Gene Mutations Are Associated with Cerebral Large-Artery Atherosclerosis.

BACKGROUND: To investigate whether one or more SAMHD1 gene mutations are associated with cerebrovascular disease in the general population using a Chinese stroke cohort. METHODS: Patients with a Chinese Han background (N = 300) diagnosed with either cerebral large-artery atherosclerosis (LAA, n = 100), cerebral small vessel disease (SVD, n = 100), or other stroke-free neurological disorders (control, n = 100) were recruited. Genomic DNA from the whole blood of each patient was isolated, and direct sequencing of the SAMHD1 gene was performed. Both wild type and mutant SAMHD1 proteins identified from the patients were expressed in E. coli and purified; then their dNTPase activities and ability to form stable tetramers were analysed in vitro. RESULTS: Three heterozygous mutations, including two missense mutations c.64C>T (P22S) and c.841G>A (p.E281K) and one splice site mutation c.696+2T>A, were identified in the LAA group with a prevalence of 3%. No mutations were found in the patients with SVD or the controls (p = 0.05). The mutant SAMHD1 proteins were functionally impaired in terms of their catalytic activity as a dNTPase and ability to assemble stable tetramers. CONCLUSIONS: Heterozygous SAMHD1 gene mutations might cause genetic predispositions that interact with other risk factors, resulting in increased vulnerability to stroke.

Induction of KIAA1199/CEMIP is associated with colon cancer phenotype and poor patient survival.

Genes induced in colon cancer provide novel candidate biomarkers of tumor phenotype and aggressiveness. We originally identified KIAA1199 (now officially called CEMIP) as a transcript highly induced in colon cancer: initially designating the transcript as Colon Cancer Secreted Protein 1. We molecularly characterized CEMIP expression both at the mRNA and protein level and found it is a secreted protein induced an average of 54-fold in colon cancer. Knockout of CEMIPreduced the ability of human colon cancer cells to form xenograft tumors in athymic mice. Tumors that did grow had increased deposition of hyaluronan, linking CEMIP participation in hyaluronan degradation to the modulation of tumor phenotype. We find CEMIP mRNA overexpression correlates with poorer patient survival. In stage III only (n = 31) or in combined stage II plus stage III colon cancer cases (n = 73), 5-year overall survival was significantly better (p = 0.004 and p = 0.0003, respectively) among patients with low CEMIP expressing tumors than those with high CEMIP expressing tumors. These results demonstrate that CEMIP directly facilitates colon tumor growth, and high CEMIP expression correlates with poor outcome in stage III and in stages II+III combined cohorts. We present CEMIP as a candidate prognostic marker for colon cancer and a potential therapeutic target.

Recessive nephrocerebellar syndrome on the Galloway-Mowat syndrome spectrum is caused by homozygous protein-truncating mutations of WDR73.

We describe a novel nephrocerebellar syndrome on the Galloway-Mowat syndrome spectrum among 30 children (ages 1.0 to 28 years) from diverse Amish demes. Children with nephrocerebellar syndrome had progressive microcephaly, visual impairment, stagnant psychomotor development, abnormal extrapyramidal movements and nephrosis. Fourteen died between ages 2.7 and 28 years, typically from renal failure. Post-mortem studies revealed (i) micrencephaly without polymicrogyria or heterotopia; (ii) atrophic cerebellar hemispheres with stunted folia, profound granule cell depletion, Bergmann gliosis, and signs of Purkinje cell deafferentation; (iii) selective striatal cholinergic interneuron loss; and (iv) optic atrophy with delamination of the lateral geniculate nuclei. Renal tissue showed focal and segmental glomerulosclerosis and extensive effacement and microvillus transformation of podocyte foot processes. Nephrocerebellar syndrome mapped to 700 kb on chromosome 15, which contained a single novel homozygous frameshift variant (WDR73 c.888delT; p.Phe296Leufs*26). WDR73 protein is expressed in human cerebral cortex, hippocampus, and cultured embryonic kidney cells. It is concentrated at mitotic microtubules and interacts with α-, ß-, and γ-tubulin, heat shock proteins 70 and 90 (HSP-70; HSP-90), and the carbamoyl phosphate synthetase 2/aspartate transcarbamylase/dihydroorotase multi-enzyme complex. Recombinant WDR73 p.Phe296Leufs*26 and p.Arg256Profs*18 proteins are truncated, unstable, and show increased interaction with α- and ß-tubulin and HSP-70/HSP-90. Fibroblasts from patients homozygous for WDR73 p.Phe296Leufs*26 proliferate poorly in primary culture and senesce early. Our data suggest that in humans, WDR73 interacts with mitotic microtubules to regulate cell cycle progression, proliferation and survival in brain and kidney. We extend the Galloway-Mowat syndrome spectrum with the first description of diencephalic and striatal neuropathology.

A new liquid chromatography/tandem mass spectrometry method for quantification of gangliosides in human plasma.

Gangliosides are a family of glycosphingolipids characterized by mono- or polysialic acid-containing oligosaccharides linked through 1,3- and 1,4-ß glycosidic bonds with subtle differences in structure that are abundantly present in the central nervous systems of many living organisms. Their cellular surface expression and physiological malfunction are believed to be pathologically implicated in considerable neurological disorders, including Alzheimer and Parkinson diseases. Recently, studies have tentatively elucidated that mental retardation or physical stagnation deteriorates as the physiological profile of gangliosides becomes progressively and distinctively abnormal during the development of these typical neurodegenerative syndromes. In this work, a reverse-phase liquid chromatography/tandem mass spectrometry (LC/MS/MS) assay using standard addition calibration for determination of GM2, GM3, GD2, and GD3 in human plasma has been developed and validated. The analytes and internal standard were extracted from human plasma using a simple protein precipitation procedure. Then the samples were analyzed by reverse-phase ultra-performance liquid chromatography (UPLC)/MS/MS interfaced to mass spectrometry with electrospray ionization using a multiple reaction monitoring mode to obtain superior sensitivity and specificity. This assay was validated for extraction recovery, calibration linearity, precision, and accuracy. Our quick and sensitive method can be applied to monitor ganglioside levels in plasma from normal people and neurodegenerative patients.

Cutaneous dyspigmentation in patients with ganglioside GM3 synthase deficiency.

Ganglioside GM3 synthase deficiency is a rare autosomal recessive metabolic disorder characterized by infantile onset of severe irritability and epilepsy, failure to thrive, developmental stagnation, and cortical blindness. Because of the lack of easily recognizable dysmorphism and specific neurologic manifestations, identification of patients with this condition is extremely challenging. Here we report on previously undescribed pigmentary abnormalities in 20 of 38 patients with GM3 synthase deficiency. All 20 of the patients showed freckle-like hyperpigmented macules, ranging in size from 2 to 5 mm in diameter and usually found bilaterally on the extremities, especially the dorsal aspects of the hands and feet. Seven of these patients also had depigmented macules and patches, especially on the face and extremities. These cutaneous changes were asymptomatic, and were not associated with the severity or particular phenotype of the neurologic disease. They became visible only after the first years of life with an increased incidence with advancing age. These distinct pigmentary features are not identified in 54 normal siblings, and may provide a useful clue in identifying patients with ganglioside metabolic disorders.

A homozygous missense mutation in HERC2 associated with global developmental delay and autism spectrum disorder.

We studied a unique phenotype of cognitive delay, autistic behavior, and gait instability segregating in three separate sibships. We initiated genome-wide mapping in two sibships using Affymetrix 10K SNP Mapping Arrays and identified a homozygous 8.2 Mb region on chromosome 15 common to five affected children. We used exome sequencing of two affected children to assess coding sequence variants within the mapped interval. Four novel homozygous exome variants were shared between the two patients; however, only two variants localized to the mapped interval on chromosome 15. A third sibship in an Ohio Amish deme narrowed the mapped interval to 2.6 Mb and excluded one of the two novel homozygous exome variants. The remaining variant, a missense change in HERC2 (c.1781C>T, p.Pro594Leu), occurs in a highly conserved proline residue within an RCC1-like functional domain. Functional studies of truncated HERC2 in adherent retinal pigment epithelium cells suggest that the p.Pro594Leu variant induces protein aggregation and leads to decreased HERC2 abundance. The phenotypic correlation with the mouse Herc1 and Herc2 mutants as well as the phenotypic overlap with Angelman syndrome provide further evidence that pathogenic changes in HERC2 are associated with nonsyndromic intellectual disability, autism, and gait disturbance. Hum Mutat 33:1639-1646, 2012. © 2012 Wiley Periodicals, Inc.

Heterozygote genotypes at rs2222823 and rs2811712 SNP loci are associated with cerebral small vessel disease in Han Chinese population.

AIMS: With developments of etiology of cerebral small vessel disease (CSVD) and genome-wide association study (GWAS) of stroke, the genetic studies of CSVD are focused on genes related to blood-brain barrier (BBB) and aging. The project aims to investigate the association between CSVD and susceptibility loci and candidate genes. METHODS: All study subjects admitted Beijing Tiantan Hospital from June 2009 to September 2010 including 197 cerebral small vessel disease patients(S), 198 large artery atherosclerosis control individuals (vascular stenotic rate ≥50% diameter reduction) (L), 200 hypertensive intracerebral hemorrhage control individuals (H) and 197 stroke-free control individuals (C). 15 SNPs in 4 genes (MYLK, AQP4, NINJ2, and INK4/ARF) were genotyped using Multiplex Snapshot assay. Each SNP was first examined between the groups S and C in different genetic models (codominant, dominant, recessive, overdominant, and log-additive). Permutation correction was used to adjust for multiple testing. The significant SNP loci were further analyzed in comparing S with L and H, respectively. Subgroup analysis was also performed for each risk-factor category. RESULTS: Among the 15 SNPs, rs2222823 and rs2811712 were found to be significantly associated with CSVD after multiple-testing adjustment. The heterozygote (A/T) of rs2222823 of MYLK has an odds ratio of 0.52 (95% CI =[0.35, 0.79], P= 0.002, adjusted P= 0.031) when compared with homozygotes. The heterozygote (C/T) of rs2811712 of INK4/ARF has an odds ratio of 1.75 (95% CI =[1.13-2.71], P= 0.004, adjusted P= 0.050). The SNP rs2222823 was significant (P= 0.035) in comparing S with H. In comparing S versus L, it is significant for the subgroups of patients without diabetes (P= 0.012) and drinking (P= 0.018). rs2811712 was significant in comparing S with L for the subgroups of patients with hyperlipidemia (P= 0.029) and drinking (P= 0.04). CONCLUSION: The heterozygotes (T/A) at the rs2222823 SNP locus of MYLK gene decreases the risk of having cerebral small vessel disease, while the heterozygotes (C/T) at the rs2811712 SNP locus of INK4/ARF gene increases the risk, suggesting that the MYLK and INK4/ARF are the associated genes of cerebral small vessel disease in Han Chinese population.

Subclinical echocardiographic abnormalities in phenotype-negative carriers of myosin-binding protein C3 gene mutation for hypertrophic cardiomyopathy.

BACKGROUND: Early diastolic myocardial tissue Doppler velocities have reported to be reduced in mutation-positive patients with hypertrophic cardiomyopathy (HCM) in some studies even in the absence of left ventricular hypertrophy (LVH). Strain is a sensitive tool in detecting early systolic abnormalities in patients with HCM. Our goal is to examine novel echocardiographic characteristics of phenotype-negative carriers for a known sarcomeric gene mutation for HCM. METHODS: We evaluated 41 consecutive subjects with a known myosin-binding protein C3 (MYBPC3) mutation (c.3330+2T>G). Subjects who were mutation positive without LVH (G+/LVH-, n = 35) were compared with healthy controls (n = 30) regarding tissue Doppler and segmental longitudinal strain measures. RESULTS: The G+/LVH- group was similar to the healthy controls with respect to chamber size, left ventricular mass index, and most diastolic filling parameters, including tissue Doppler-derived early diastolic annular velocities. Global longitudinal strain was similar for both groups (20.3 ± 2.1 vs 19.8 ± 1.8, P = .36), although regional segment analysis showed a notable reduction in the basal septum (16.8 ± 3.1 vs 19.0 ± 4.0%, P = .02) and increase in the basal posterior (22.5 ± 5.2 vs 17.9 ± 5.2, P = .001) as well as mid posterior (21.8 ± 4.7 vs 18.2 ± 3.0, P = .001) walls. CONCLUSIONS: In our cohort of phenotype-negative carriers of a specific MYBPC3 mutation, there were minimal differences in conventional 2-dimensional, Doppler, and speckle-tracking-derived parameters of systolic and diastolic function compared with that of healthy subjects. The presence of regional alterations in strain indicative of the presence of underlying subclinical disease requires further validation.

Mutations in U4atac snRNA, a component of the minor spliceosome, in the developmental disorder MOPD I.

Small nuclear RNAs (snRNAs) are essential factors in messenger RNA splicing. By means of homozygosity mapping and deep sequencing, we show that a gene encoding U4atac snRNA, a component of the minor U12-dependent spliceosome, is mutated in individuals with microcephalic osteodysplastic primordial dwarfism type I (MOPD I), a severe developmental disorder characterized by extreme intrauterine growth retardation and multiple organ abnormalities. Functional assays showed that mutations (30G>A, 51G>A, 55G>A, and 111G>A) associated with MOPD I cause defective U12-dependent splicing. Endogenous U12-dependent but not U2-dependent introns were found to be poorly spliced in MOPD I patient fibroblast cells. The introduction of wild-type U4atac snRNA into MOPD I cells enhanced U12-dependent splicing. These results illustrate the critical role of minor intron splicing in human development.

Homozygous mutation in SAMHD1 gene causes cerebral vasculopathy and early onset stroke.

We describe an autosomal recessive condition characterized with cerebral vasculopathy and early onset of stroke in 14 individuals in Old Order Amish. The phenotype of the condition was highly heterogeneous, ranging from severe developmental disability to normal schooling. Cerebral vasculopathy was a major hallmark of the condition with a common theme of multifocal stenoses and aneurysms in large arteries, accompanied by chronic ischemic changes, moyamoya morphology, and evidence of prior acute infarction and hemorrhage. Early signs of the disease included mild intrauterine growth restriction, infantile hypotonia, and irritability, followed by failure to thrive and short stature. Acrocyanosis, Raynaud's phenomenon, chilblain lesions, low-pitch hoarse voice, glaucoma, migraine headache, and arthritis were frequently observed. The early onset or recurrence of strokes secondary to cerebral vasculopathy seems to always be associated with poor outcomes. The elevated erythrocyte sedimentation rate (ESR), IgG, neopterin, and TNF-α found in these patients suggested an immune disorder. Through genomewide homozygosity mapping, we localized the disease gene to chromosome (Chr) 20q11.22-q12. Candidate gene sequencing identified a homozygous mutation, c.1411-2A > G, in the SAMHD1 gene, being associated with this condition. The mutation appeared at the splice-acceptor site of intron 12, resulted in the skipping of exon 13, and gave rise to an aberrant protein with in-frame deletion of 31 amino acids. Immunoblotting analysis showed lack of mutant SAMHD1 protein expression in affected cell lines. The function of SAMHD1 remains unclear, but the inflammatory vasculopathies of the brain found in the patients with SAMHD1 mutation indicate its important roles in immunoregulation and cerebral vascular hemeostasis.

Homozygous frameshift mutation in TMCO1 causes a syndrome with craniofacial dysmorphism, skeletal anomalies, and mental retardation.

We identified an autosomal recessive condition in 11 individuals in the Old Order Amish of northeastern Ohio. The syndrome was characterized by distinctive craniofacial dysmorphism, skeletal anomalies, and mental retardation. The typical craniofacial dysmorphism included brachycephaly, highly arched bushy eyebrows, synophrys, long eyelashes, low-set ears, microdontism of primary teeth, and generalized gingival hyperplasia, whereas Sprengel deformity of scapula, fusion of spine, rib abnormities, pectus excavatum, and pes planus represented skeletal anomalies. The genome-wide homozygosity mapping using six affected individuals localized the disease gene to a 3.3-Mb region on chromosome 1q23.3-q24.1. Candidate gene sequencing identified a homozygous frameshift mutation, c.139_140delAG, in the transmembrane and coiled-coil domains 1 (TMCO1) gene, as the pathogenic change in all affected members of the extended pedigree. This mutation is predicted to result in a severely truncated protein (p.Ser47Ter) of only one-fourth the original length. The TMCO1 gene product is a member of DUF841 superfamily of several eukaryotic proteins with unknown function. The gene has highly conserved amino acid sequence and is universally expressed in all human tissues examined. The high degree of conservation and the ubiquitous expression pattern in human adult and fetal tissues suggest a critical role for TMCO1. This report shows a TMCO1 sequence variant being associated with a genetic disorder in human. We propose "TMCO1 defect syndrome" as the name of this condition.