An expansion of phenotype: novel homozygous variant in the MED17 identified in patients with progressive microcephaly and global developmental delay.

Global developmental delay (GDD) is a lifelong disability that affects 1-3% of the population around the globe. It is phenotypically variable and highly heterogeneous in terms of the underlying genetics. Patients with GDD are intellectually disabled (ID) manifesting cognitive impairment and deficient adaptive behavior. Here, we investigated a two-looped consanguineous family segregating severe ID, seizure, and progressive microcephaly. Magnetic resonance imaging (MRI) of the brain showed mild brain atrophy and myelination defect. Whole exome sequencing (WES) was performed on the DNA samples of two patients and a novel homozygous missense variant (Chr11:g0.93528085; NM_004268.5_c.871T > C; p. Trp291Gly) was identified in the MED17 gene. Sanger sequencing revealed that the identified variant is heterozygous in both parents and healthy siblings. This variant is conserved among different species, causes a non-conserved amino acid change, and is predicted deleterious by various in silico tools. The variant is not reported in population variant databases. MED17 (OMIM: 613668) encodes for the mediator of RNA polymerase II transcription complex subunit 17. Structure modeling of MED17 protein revealed that Trp291 is involved in different inter-helical interactions, providing structural stability. Replacement of Trp291Gly, a less hydrophobic amino acid loses the inter-helical interaction leading to a perturb variant of MED17 protein.

Clinical and genetic characterization of patients segregating variants in KPTN, MINPP1, NGLY1, AP4B1, and SON underlying neurodevelopmental disorders: Genetic and phenotypic expansion.

Neurodevelopmental disorders (NDDs) are heterogeneous genetic conditions of the central nervous system (CNS). Primary phenotypes of NDDs include epilepsy, loss of developmental skills, abnormal movements, muscle weakness, ocular anomalies, hearing problems, and macro- or microcephaly. NDDs occur due to variants in genes encoding proteins involved in the structure and function of CNS, thus interrupting its normal physiological role. In the study presented here, four consanguineous families (A-D), with members showing neurodevelopmental symptoms, were recruited for clinical and genetic characterization of the phenotypes. Clinical examinations, including Seguin Form Board Test (SFBT), Vineland Social Maturity Scale (VSMS), brain Magnetic Resonance Imaging (MRI), Electroencephalogram (EEG), Electromyography (EMG), Nerve Conduction Velocity (NCV), and Magnetic Resonance Spectroscopy, were employed to characterize the disease phenotypes. Whole exome sequencing (WES) followed by Sanger sequencing was employed to search for the genetic basis of the neurological symptoms observed in four families (A-D). Two of these families (A, B) were of Saudi Arabian origin, and two others (C, D) were of Pakistan origin. Two homozygous missense (KPTN: NM_007059.4:c.301T>G: NP_008990.2; p.(Phe101Val) and MINPP1:NM_001178118.2:c.1202G>A: NP_001171588.1; p.(Arg401Gln)) variants in families A and B, respectively, and two homozygous nonsense (NGLY1:NM_018297.3:c.1534_1541dup: NP_060767.2; p.(Ser515LysfsTer51) and AP4B1:NM_001253852:c.1668G>A: NP_001240781.1; p.(Trp556X)) variants in families C and D, respectively, were identified. Interestingly, additional heterozygous nonsense variant in SON: NM_138927.2: c.5753_5756del: NP_620305.3; p.(Val1918GlufsTer87) and a homozygous variant in FLG (FLG: NM_002016.2:c.7339C>T: NP_002007.1; p.(Arg2447X) were detected in families A and D, respectively. Further, we determined the deleteriousness of each variant through computational approaches. The present study expands the phenotypic and genetic spectrum of NDD-associated genes (KPTN, MINPP1, NGLY1, and AP4B1). Moreover, additional nonsense variants (SON: c.5753_5756del and FLG: c.7339C>T) identified in two families segregating with the phenotype might explain the phenotypic variability and severity in our patients.

A Novel Frameshift Mutation in the ITGB3 Gene Leading to Glanzmann's Thrombasthenia in a Saudi Arabian Family.

Glanzmann's thrombasthenia (GT) is an autosomal recessive congenital bleeding disorder of platelet aggregation. Mutations in ITGA2B and ITGB3 genes result in quantitative and/or qualitative abnormalities of the glycoprotein receptor complex IIb/IIIa (integrin αIIbß3), which in turn impairs platelet aggregation and lead to GT. In this study, whole genome single nucleotide polymorphism (SNP) genotyping as well as whole exome sequencing was performed in a large family segregating GT. Analysis of the genotypes localized the disease region to chromosome 17q21.2-q21.3. Filtration of whole exome data and candidate variants prioritization identified a pathogenic variant in the ITGB3 gene. The single nucleotide deletion variant (c.2113delC) in exon 13 of the ITGB3 gene is predicted to cause a frameshift and absence of vital C-terminal domains including the transmembrane helix and the cytoplasmic domain. Clinical variability of the bleeding phenotype in affected individuals with the same mutation suggests that other genetic and nongenetic factors are responsible for determining GT features.

Frameshift variant in MITF gene in a large family with Waardenburg syndrome type II and a co-segregation of a C2orf74 variant.

Waardenburg syndrome (WS) is a hereditary disorder affecting the auditory system and pigmentation of hair, eyes, and skin. Different variants of the disease exist with the involvement of mutation in six genes. The aim of the study is to identify the genetic defects underlying Waardenburg syndrome in a large family with multiple affected individuals. Here, in this study, we recruited a large family with eleven affected individuals segregating WS type 2. We performed whole genome SNP genotyping, whole exome sequencing and segregation analysis using Sanger approach. Whole genome SNP genotyping, whole exome sequencing followed by Sanger validation of variants of interest identified a novel single nucleotide deletion mutation (c.965delA) in the MITF gene. Moreover, a rare heterozygous, missense damaging variant (c.101T>G; p.Val34Gly) in the C2orf74 has also been identified. The C2orf74 is an uncharacterized gene present in the linked region detected by DominantMapper. Variants in MITF and C2orf74 follows autosomal dominant segregation with the phenotype, however, the variant in C2orf74 is incompletely penetrant. We proposed a digenic inheritance of variants as an underlying cause of WS2 in this family.

Genetic variations in autoimmune genes and VKH disease.

PURPOSE: Vogt-Koyanagi-Harada (VKH) disease is a rare autoimmune disease. The autoimmune response in VKH disease is against the melanin-producing cells; therefore, in affected individuals melanocyte-containing organs manifest disease symptoms including eyes, ears, skin and nervous system. VKH is a multifactorial disease, and the precise cause of the VKH disease is unknown. Studies have suggested that both environmental and genetic factors are responsible for the VKH disease. In this review, the authors have collected all the available literature on the genetics of VKH to their knowledge and discussed the role of genetic variants in causing VKH disease. METHODS: An extensive literature search was performed in order to review all the published studies regarding VKH clinical phenotyping and genetic variants in VKH disease. Medline, PubMed, Cochrane library, and Scopus was searched using combination of keywords. RESULTS: It was found that variants in HLA genes, IL-12b, TNFSF4, and miR-20-5p genes are significantly associated with VKH; however, variants in genes ATG10, TNIP1 and CLEC16A did not achieve significant genome-wide association threshold. Moreover, polymorphisms in TNIP1 and CLEC16A play a protective role against VKH. CONCLUSION: The authors conclude that increased sample size and a more homogeneous VKH patient population may reveal a significant association of variants in ATG10, TNIP1 and CLEC16A genes with VKH disease.

A novel missense variant in the RASGRP2 gene in patients with moderate to severe bleeding disorder.

Inherited platelet function disorder-18 (IPD-18) is a relatively new non-syndromic autosomal recessive bleeding disorder. It is characterized by deficient or dysfunctional CalDAG-GEFI protein. The distinctive feature of the disease is impaired platelet aggregation in response to multiple physiologic agonists. We here report a family with a platelet-type bleeding disorder and a novel mutation in the RASGRP2 gene. The overall bleeding score for the affected individuals was 15 and 12. Based on the initial diagnosis of Glanzmann thrombasthenia, targeted sequencing of integrin subunit alpha 2b and integrin subunit beta 3 encoding genes ITGA2B and ITGB3 were carried out in both affected members of a family. Sequence alignment failed to identify the disease-causing variant(s) in both genes. Therefore, whole exome sequencing in one affected individual was performed. Data analysis detected a novel homozygous missense variant (c.956C>T; p.Pro319Leu) in the exon 9 of the RASGRP2 gene. Five additional individuals of a family including both parents, an affected individual and two asymptomatic individuals were Sanger sequenced for the variant (c.956C>T). The variant segregates in the family in an autosomal recessive manner. Several in silico tools predicted the variant as pathogenic. Protein modeling studies suggest that the mutation (p.Pro319Leu) cause a conformational change in the loop structure of the RasGEF domain of the CalDAG-GEFI protein. Reported variants in the RasGEF domain impair expression and/or nucleotide exchange activity of CalDAG-GEFI protein and thus inhibit the activation of Rap1 protein required for platelet adhesion and hemostatic plug formation.

A homozygous missense variant in the homeobox domain of the NKX6-2 results in progressive spastic ataxia type 8 associated with lower limb weakness and neurological manifestations.

BACKGROUND: Progressive spastic ataxia is a heterogeneous disorder characterized by cerebellar ataxia and limb spasticity associated with other severe neurological complications. Spastic ataxia is classified into pure and complex types, inherited in both an autosomal recessive and autosomal dominant manner. It is caused by pathogenic variants in at least eight different genes, including NKX6-2 (MIM 607063) located on chromosome 10q26.3. The present study aimed to identify the genetic variant(s) underlying progressive spastic ataxia and to establish the genotype-phenotype correlation. METHODS: We collected a large consanguineous family having four affected individuals segregating progressive spastic ataxia in an autosomal recessive manner. To investigate the molecular cause of the disease, genomic DNA of three affected individuals underwent whole exome sequencing. RESULTS: All of the affected individuals showed progressive clinical features such as spastic ataxia, lower limb weakness and other mild neurological abnormalities. Whole exome sequencing data were analyzed using different filters. Filtering of rare and shared homozygous variants revealed a novel homozygous missense variant (c.545C>T; p.Ala182Val) in a highly conserved homeobox domain of the NKX6-2 protein. CONCLUSIONS: The findings of the present study add a novel variant to the NKX6-2 mutation spectrum and provide evidence that homozygous variants in the NKX6-2 cause progressive spastic ataxia associated with other abnormalities.

Novel homozygous loss-of-function mutations in RP1 and RP1L1 genes in retinitis pigmentosa patients.

Background: Retinitis pigmentosa (RP) is a heterogeneous group of ocular dystrophy. It is challenging to identify the underlying genetic defect in individuals with RP due to huge genetic heterogeneity. This study was designed to delineate the genetic defect(s) underlying RP in extended Saudi families and to describe the possible disease mechanism.Materials and Methods: Fundus photography and a high definition optical coherence tomography (HD-OCT) were performed in order to detect the earlier stages of macular degeneration. Genomic DNA was extracted followed by genome-wide SNP genotyping and whole exome sequencing (WES). Exome data was filtered to identify the genetic variant(s) of interest.Results: Clinical examination showed that affected individuals manifest key features of RP. The fundus exam shows pale optic disc and bone spicules at the periphery. OCT shows macular degeneration as early as at the age of 4 years. Whole genome scan by SNPs identified multiple homozygous regions. WES identified a 10 bps novel insertion mutation (c.3544_3545insAGAAAAGCTG; p.Ala1182fs) in the RP1 gene in both affected individuals of family A. Affected individual from family B showed a large insertion of 48 nucleotides in the coding part of the RP1L1 gene (c.3955_3956insGGACTAAAGTAATAGAAGGGCTGCAAGAAGAGAGGGTGCAGTTAGAGG; p.Ala1319fs). Sanger sequencing validates the autosomal recessive inheritance of the mutations.Conclusion: The results strongly suggest that the insertion mutations in the RP1 and RP1L1 genes are responsible for the retinal phenotype in affected individuals from two families. Heterozygous individuals are asymptomatic carriers. We propose that the protective allele in other homozygous regions in heterozygous carriers contribute to the phenotypic variability in asymptomatic individuals.

KMT2C, a histone methyltransferase, is mutated in a family segregating non-syndromic primary failure of tooth eruption.

Primary failure of tooth eruption (PFE) is a rare odontogenic defect and is characterized by failure of eruption of one or more permanent teeth. The aim of the study is to identify the genetic defect in a family with seven affected individuals segregating autosomal dominant non-syndromic PFE. Whole genome single-nucleotide polymorphism (SNP) genotyping was performed. SNP genotypes were analysed by DominantMapper and multiple shared haplotypes were detected on different chromosomes. Four individuals, including three affected, were exome sequenced. Variants were annotated and data were analysed while considering candidate chromosomal regions. Initial analysis of variants obtained by whole exome sequencing identified damaging variants in C15orf40, EPB41L4A, TMEM232, KMT2C, and FBXW10 genes. Sanger sequencing of all family members confirmed segregation of splice acceptor site variant (c.1013-2 A > G) in the KMT2C gene with the phenotype. KMT2C is considered as a potential candidate gene based on segregation analysis, the absence of variant in the variation databases, the presence of variant in the shared identical by descent (IBD) region and in silico pathogenicity prediction. KMT2C is a histone methyltransferase and recently the role of another member of this family (KMT2D) has been implicated in tooth development. Moreover, protein structures of KMT2C and KMT2D are highly similar. In conclusion, we have identified that the KMT2C gene mutation causes familial non-syndromic PFE. These findings suggest the involvement of KMT2C in the physiological eruption of permanent teeth.

Exome sequencing revealed a novel loss-of-function variant in the GLI3 transcriptional activator 2 domain underlies nonsyndromic postaxial polydactyly.

BACKGROUND: Polydactyly is a common genetic limb deformity characterized by the presence of extra fingers or toes. This anomaly may occur in isolation (nonsyndromic) or as part of a syndrome. The disease is broadly divided into preaxial polydactyly (PPD; duplication of thumb), mesoaxial polydactyly (complex polydactyly), and postaxial polydactyly (PAP: duplication of the fifth finger). The extra digits may be present in one or both the limbs. Heterozygous variants in the GLI3, ZRS/SHH, and PITX1 have been associated with autosomal dominant polydactyly, while homozygous variants in the ZNF141, IQCE, GLI1, and FAM92A have been associated with autosomal recessive polydactyly. Pathogenic mutations in the GLI3 gene (glioma-associated oncogene family zinc finger 3) have been associated with both nonsyndromic and syndromic polydactyly. METHODS: Here, we report an extended five generation kindred having 12 affected individuals exhibiting nonsyndromic postaxial polydactyly type A condition. Whole-exome sequencing followed by variant prioritization, bioinformatic studies, Sanger validation, and segregation analysis was performed. RESULTS: Using exome sequencing in the three affected individuals, we identified a novel heterozygous frameshift variant (c.3567_3568insG; p.Ala1190Glyfs*57) in the transcriptional activator (TA2) domain of the GLI3 encoding gene. CONCLUSION: To the best of our knowledge, the present study reports on the first familial case of nonsyndromic postaxial polydactyly due to the GLI3 variant in Pakistani population. Our study also demonstrated the important role of GLI3 in causing nonsyndromic postaxial polydactyly.

A Heterozygous Mutation in the Triple Helical Region of the Alpha 1 (II) Chain of the COL2A1 Protein Causes Non-Lethal Spondyloepiphyseal Dysplasia Congenita.

Objective:Heterozygous pathogenic variants in the COL2A1 gene result in several clinical features including impaired skeletal growth, ocular and otolaryngological abnormalities. Missense mutations in the triple helical region of the COL2A1 protein have been associated with lethal spondyloepiphyseal dysplasia (SED). In this study, we aimed to identify the underlying cause of a case of SED congenita (SEDC) in a 27-month-old child. Materials and Methods: A patient who was diagnosed initially with osteochondrodysplasia underwent a detailed clinical and radiological examination to obtain a conclusive diagnosis. The patient did not show any clinical features of hypochondrogenesis. Whole exome sequencing of the COL2A1 gene was carried out to identify the underlying genetic cause of the disorder. Results: Variant annotation and filtration detected a heterozygous missense mutation c.1357G>A (p.G453S) in the exon 21 of the COL2A1 gene of the proband which was confirmed by Sanger sequencing. Neither parent carried the mvariant suggesting this was a new mutation. Conclusion: The COL2A1 mutation (c.1357G>A), identified in this case, results in more mild phenotype than other missense mutations in exon 21 which are known to cause lethal hypochondrogenesis. We showed, for the first time, that a missense mutation (p.G453S) in the triple helical region of the alpha 1 (II) chain of the COL2A1 protein underlies SEDC and is not always lethal.

Whole genome genotyping mapped regions on chromosome 2 and 18 in a family segregating Waardenburg syndrome type II.

OBJECTIVES: Waardenburg syndrome is a rare genetic disorder. It is characterized by sensorineural hearing impairment and pigment defects of the skin, hair and iris. In some cases abnormalities in the tissues derived from neural crest have also been reported. Mutations in several genes have been reported as an underlying cause of Waardenburg syndrome. Objective of this study is to identify the chromosomal region(s) associated with Waardenburg syndrome in an extended Saudi family. METHODS: Genomic DNA was extracted from fifteen individuals of a Saudi family segregating Waardenburg syndrome. Whole genome SNP genotyping was performed to identify common identity by descent chromosomal region(s) shared by affected individuals. RESULTS: Pedigree analysis confirm autosomal dominant inheritance of Waardenburg syndrome type II in a family. Whole genome SNP genotypes were analyzed using AutoSNPa and DominantMapper tools. Shared identity by descent chromosomal regions were identified on chromosome 2 and chromosome 18. Regions were checked for known Waardenburg syndrome genes. No known gene is present in both regions. CONCLUSIONS: In summary, we identified novel chromosomal regions associated with Waardenburg syndrome type II in a Saudi family. Deep sequencing of a complete candidate regions are required to identify the gene underlying Waardenburg syndrome in this family.

Whole exome sequencing identification of a novel insertion mutation in the phospholipase C ε­1 gene in a family with steroid resistant inherited nephrotic syndrome.

Nephrotic syndrome (NS) represents a heterogeneous group of kidney disorders characterized by excessive proteinuria, hypoalbuminemia and edema. Defects in the filtration barrier of the glomeruli results in the development of NS. The genetic cause of NS remains to be fully elucidated. However, previous studies based on positional cloning of genes mutated in NS have provided limited insight into the pathogenesis of this disease. Mutations in phospholipase C ε­1 (PLCE1) have been reported as a cause of early onset NS characterized by histology of diffuse mesangial sclerosis. In the present study, the underlying cause of NS in a consanguineous family was identified. Clinical and molecular aspects of a consanguineous Saudi family comprised of five individuals with steroid resistant NS were examined. Seven healthy individuals from the same family were also studied. Whole exome sequencing (WES) was performed to detect the genetic defect underlying NS. WES identified a homozygous novel insertion mutation (c.6272_6273insT) in the PLCE1 gene. Pedigree and segregation analysis confirmed an autosomal recessive inheritance pattern. This mutation may result in a bi­allelic loss of the C­terminal Ras­associating domain in PLCE1 that results in NS. The present study expanded the mutational spectrum of PLCE1 in NS. In addition, the present study provided further evidence that supports the important involvement of PLCE1 in the physiological function of the glomerular filtration barrier.

Whole exome sequencing identified a novel single base pair insertion mutation in the EYS gene in a six generation family with retinitis pigmentosa.

Retinitis pigmentosa (RP) is a group of inherited progressive retinal dystrophies (RD) and is characterized by photoreceptor degeneration. RP is clinically and genetically heterogeneous disorder. More than 70 genes are known and, thus, identification of causative genes and mutations in known genes is challenging. This study was designed to identify the underlying genetic defect in a large extended Saudi family with multiple RP affected members. Fundus photography, Optical Coherence Tomography (OCT) and visual field perimetry were performed for affected individuals. Whole exome sequencing was used to detect the underlying genetic defect in a large family with 12 affected individuals showing autosomal recessive isolated RP. WES data analysis identified a novel insertion mutation in the EYS (eyes shut homolog) gene (c.910_911insT; p.Trp304LeufsTer8). Sanger sequencing validates the variant discovered through exome in all 12 affected individuals and showed that this mutation is segregating with RP phenotype in an autosomal recessive manner in 51 individuals of the family tested here. Our study expands the mutation spectrum of EYS gene in RP patients and extends the body of evidence that supports the importance of EYS gene in eye development.

Whole genome SNP genotyping in a family segregating developmental dysplasia of the hip detected runs of homozygosity on chromosomes 15q13.3 and 19p13.2.

Developmental dysplasia of the hip (DDH) is one of the most prevalent developmental orthopedic diseases worldwide. DDH is a spectrum of anatomical abnormalities of the hip joint and is characterized by premature arthritis in later life. Sporadic cases have been reported more frequently; however, some studies have reported families segregating DDH. Studies have suggested that the genetic factors play a significant role in the development of DDH. In order to detect genetic defect underlying DDH, we performed Sanger sequencing of all DDH associated genes, whole genome SNP genotyping and exome sequencing in a Saudi family with four individuals having DDH. Sanger sequencing of all known genes did not identify any pathogenic variant. Genotype data analysis using HomozygosityMapper identified shared homozygous regions on chromosome 15q13.3 and chromosome 19p13.2 flanked by rs17228178-rs1534200 and rs466123-rs2112461, respectively. These data were also analyzed by cnvpartition software for identification of DDH associated copy number variations (CNV). A shared copy number gain of approximately 15 kb on chr6p21.32 (chr6:33 053 906-33 069 893) was discovered in all affected individuals. Partial gain of this region has also been found in unaffected sibling of this family. Exome data did not reveal any candidate sequence variant. Whole genome sequencing is required to identify deep intronic variants in the shared homozygous regions. Identification of genetic variants involved in pathogenesis of DDH may open up interesting perspectives into the function of the gene(s) in hip joint development.

Exome sequencing identified rare variants in genes HSPG2 and ATP2B4 in a family segregating developmental dysplasia of the hip.

BACKGROUND: Developmental dysplasia of the hip (DDH) is a common pathological condition of the musculoskeletal system in infants which results in a congenital and developmental malformation of the hip joint. DDH is a spectrum of pathologies affecting the infant hip ranging from asymptomatic subtle radiographic signs through mild instability to frank dislocations with acetabular dysplasia. A Saudi family with three affected individuals with DDH was identified and genetic analysis was performed to detect the possible genetic defect(s) underlying DDH in the affected members of the family. METHODS: We performed whole genome genotyping using Illumina HumanOmni 2.5 M array and whole exome sequencing (WES) using Nextera Rapid capture kit and Illumina NextSeq500 instrument in four individuals of a family with DDH. RESULTS: SNP data analysis did not identify any runs of homozygosity and copy number variations. Identity-by-descent (IBD) analysis on whole genome genotyping data identified a shared haplotypes on chromosome 1 in affected individuals. An analysis of the WES data identified rare heterozygous variants in HSPG2 and ATP2B4 genes in the affected individuals. Multiple prediction software predicted that the variants identified are damaging. Moreover, in silico analysis showed that HSPG2 regulates ATP2B4 expression using a variety of transcription factors. CONCLUSION: Our results indicate that there might be a functional epistatic interaction between HSPG2 and ATP2B4, and DDH in the family studied is due to a combined effect of both variants. These variants are also present in the asymptomatic mother suggesting that the variants in HSPG2 and ATP2B4 are incompletely penetrant. This study provides the first evidence of digenic inheritance of DDH in a family and extends the spectrum of genetic heterogeneity in this human disorder.

A novel splice-site mutation in the ASPM gene underlies autosomal recessive primary microcephaly.

BACKGROUND: Autosomal recessive primary microcephaly (MCPH) is a clinically and genetically heterogeneous disorder. Patients with MCPH exhibit reduced occipito-frontal head circumference and non-progressive intellectual disability. To date, 17 genes have been known as an underlying cause of MCPH in humans. ASPM (abnormal spindle-like, microcephaly associated) is the most commonly mutated MCPH gene. OBJECTIVE: Identify the genetic defect underlying MCPH in a Saudi family. DESIGN: A cross-sectional clinical genetic study of a Saudi family. SETTING: Madinah Maternity and Children Hospital and Centre for Genetics and Inherited Diseases, Taibah University. PATIENTS AND METHODS: A molecular analysis was carried out on DNA samples from 10 individuals of a Saudi family segregating MCPH. DNA was isolated from the peripheral blood of 10 individuals, including 2 patients, and whole exome sequencing was performed using the Nextera Rapid Capture kit and NextSeq500 instrument. VariantStudio was used to filter and prioritize variants. MAIN OUTCOME MEASURE(S): Detection of mutation in the ASPM gene in a family segregating autoso- mal recessive primary microcephaly. RESULTS: A novel homozygous splice-site variant (c.3742-1G > C) in the ASPM gene was identified. The variant is predicted to have an effect on splicing. Human Splice Finder, an in silico tool, predicted skipping of exon 16 due to this variant. CONCLUSION: Skipping of exon 16 may change the order and number of IQ motifs in the ASPM protein leading to typical MCPH phenotype. LIMITATIONS: Single family study.

High-resolution SNP genotyping platform identified recurrent and novel CNVs in autism multiplex families.

Single nucleotide polymorphisms (SNPs)-based genotyping using microarray platform is now frequently used to detect copy number variants (CNVs) in the human genome. Here, we report CNVs identified using Illumina Human Omni 2.5M oligonucleotide microarrays in 11 multiplex families with autism spectrum disorder (ASD) referred to Autism Research and Treatment Center (ART) and Madinah Maternity and Children Hospital (MMCH). Of the 11 families, 22 patients with ASD (all males) and their parents, were recruited for the present study. In total, 43 individuals were genotyped with high-resolution array. Abnormal microarray results were seen in all 22 patients with ASD. A total of 17 shared CNVs were selected for further analysis. Out of these 17 CNVs, we discovered one novel CNV, previously not described, and 16 recurrent CNVs that overlap with the genomic imbalances defined in the autism database, autism chromosome rearrangement database and database of genomic variants. Recurrent CNVs include 11 common and 5 rare CNVs. All rare CNVs are duplications except a 16-kb deletion on chr2q36.3. Rare gain of copy numbers includes a 2-kb duplication on chr9q21.13, overlapping duplications of 107kb and 181kb on chrXp22.33 in 2 different families and a 10-kb duplication on chr18q21.13. A novel loss of copy number on chr3q23 was found in four ASD cases. This CNV results in deletion of intron 2 of calsyntenin 2 (CLSTN2) encoding synaptic protein calsyntenin 2. CLSTN2 is expressed exclusively in the brain, with high levels occurring in cortical gamma-aminobutyric acid (GABA)ergic interneurons and in medial temporal lobe regions. These results verify the diagnostic relevance of genome-wide small common and rare CNVs and provide further evidence of the high diagnostic yield of microarray for genetic testing in children with ASD.

Sequence analysis of four vitamin D family genes (VDR, CYP24A1, CYP27B1 and CYP2R1) in Vogt-Koyanagi-Harada (VKH) patients: identification of a potentially pathogenic variant in CYP2R1.

BACKGROUND: VKH is a rare autoimmune disease. Decreased level of vitamin D has recently been found to be involved in the pathogenesis of Vogt-Koyanagi-Harada (VKH) disease. This study was designed to screen the vitamin D pathway genes for pathogenic mutations, if any, in VKH patients. METHODS: Genomic DNA was extracted from blood samples collected from patients with VKH disease and healthy controls. Entire coding region, exon-intron junctions of four genes were sequenced in DNA from 39 Saudi VKH patients and 50 ethnically matched healthy individuals. All patients and controls were unrelated. RESULTS: Vitamin D levels in VKH patients were found either insufficient (21-29 ng/mL) or deficient (<20 ng/mL). Sequencing analysis of the VDR, CYP24A1, CYP27B1 and CYP2R1 detected twelve nucleotide changes in these genes in our cohort of 39 patients; 4 of which were non-coding, 6 were synonymous coding and 2 were non-synonymous coding sequence changes. All synonymous coding variants were benign polymorphisms with no apparent clinical significance. A non-synonymous coding sequence variant (c.2 T > C; p.1Met?) found in VDR is an initiation coding change and was detected in control individuals as well, while another variant (c.852G > A; p.284 M > I) found in CYP2R1 is predicted to be disease causing by mutationtaster software. This potentially pathogenic variant was found in 17 out of 39 VKH patients. CONCLUSIONS: Screening of four Vitamin D pathway genes in 39 VKH patients shows that a potentially pathogenic sequence variant in CYP2R1 may cause VKH in a subset of patients. These findings support the previous observation that low vitamin D levels might play a role in VKH pathogenesis and mutations in genes involved in vitamin D anabolism and catabolism might be of importance in VKH pathobiology.