A loss of function mutation in the COL9A2 gene causes autosomal recessive Stickler syndrome

Stickler syndrome is characterized by ocular, auditory, skeletal, and orofacial abnormalities. We describe a family with autosomal recessive Stickler syndrome. The main clinical findings consisted of high myopia, vitreoretinal degeneration, retinal detachment, hearing loss, and short stature. Affected family members were found to have a homozygous loss‐of‐function mutation in COL9A2, c.843_c.846 + 4del8. A family with autosomal recessive Stickler syndrome was previously described and found to have a homozygous loss‐of‐function mutation in COL9A1. COL9A1, COL9A2, and COL9A3 code for collagen IX. All three collagen IX α chains, α1, α2, and α3, are needed for formation of functional collagen IX molecule. In dogs, two causative loci have been identified in autosomal recessive oculoskeletal dysplasia. This dysplasia resembles Stickler syndrome. Recently, homozygous loss‐of‐function mutations in COL9A2 and COL9A3 were found to co‐segregate with the loci. Together the data from the present study and the previous studies suggest that loss‐of‐function mutations in any of the collagen IX genes can cause autosomal recessive Stickler syndrome. © 2011 Wiley‐Liss, Inc.     

Homozygous variants in MAPRE2 and CDON in individual with skin folds,growth delay,retinal coloboma,and pyloric stenosis

Cases with multiple molecular diagnoses are challenging to diagnose clinically, yet may be resolved by unbiased exome sequencing analysis. We report an infant with developmental delay, severe growth delay, dysmorphic features, and multiple congenital anomalies including retinal coloboma, congenital pyloric stenosis, and circumferential skin creases. Exome sequencing identified a homozygous missense variant in MAPRE2 and a homozygous stopgain (nonsense) variant in CDON. Variants in MAPRE2, encoding a regulator of microtubule dynamics, lead to congenital symmetric circumferential skin creases type 2, with associated dysmorphism, small growth parameters, and congenital cardiac and genital anomalies. Monoallelic variants in CDON, encoding a coreceptor for sonic hedgehog, have been associated with autosomal dominant pituitary stalk interruption syndrome and holoprosencephaly. Cdon?/? mice have multiple eye defects including coloboma, consistent with the observed human phenotype. Thus, the complex phenotypic presentation of the infant may potentially be attributed to a dual molecular diagnosis. Furthermore, we present CDON as a candidate gene for coloboma formation in addition to the known holoprosencephaly phenotype, and propose to expand the allelic spectrum of CDON to variants associated with autosomal recessive inheritance in addition to dominant inheritance.     

Robinow syndrome in an extremely preterm infant: Novel homozygous ROR2 variant detected by rapid exome sequencing

An extremely preterm infant presented with clinical and radiological features of Robinow syndrome including butterfly vertebrae, posterior rib fusion, brachydactyly, nail hypoplasia, and retromicrognathia resulting in difficult endotracheal intubation in the intensive care setting. Rapid trio exome sequencing detected a novel homozygous likely pathogenic missense variant in the ROR2 gene, NM_004560.3:c.950A>G, p.(Tyr317Cys), for which both parents were heterozygous carriers. In-silico protein modeling predicted a deleterious effect on the function of the protein. We report an extreme premature infant with novel homozygous likely pathogenic variant in the ROR2 gene consistent with autosomal recessive Robinow syndrome. This case expands the phenotypic and genotypic spectrum of this disorder and highlights the benefit of performing rapid exome sequencing early during evaluation to aid in patient management and providing accurate genetic counseling to families.     

Exome-wide copy number variation analysis identifies a COL9A1 in frame deletion that is associated with hearing loss

Pathogenic variants in COL9A1 are primarily associated with autosomal recessive Stickler syndrome. Patients with COL9A1-associated Stickler syndrome (STL) present hearing loss (HL), ophthalmic manifestations and skeletal abnormalities. However, the clinical spectrum of patients with COL9A1 variants can also include multiple epiphyseal dysplasia, as well as non-syndromic HL that was observed in one previously reported proband. Exome sequencing was performed on the genomic DNA of an Iranian patient and his affected brother who both report non-syndromic HL. A 44.6 kb homozygous in-frame deletion spanning exons 6 to 33 of COL9A1 was detected via exome-based copy number variation analysis. The deleted exons were confirmed by PCR in the patient and his affected brother, who both have non-syndromic HL. Segregation analysis via qPCR confirmed the parents as heterozygous deletion carriers. Breakpoint analysis mapped the homozygous deletion spanning introns 5 to 33 (g.70,948,188_70,997,277del, NM_001851.4(COL9A1):c.697–3754_2112+769del, p.(Phe233_Ser704del), with an additional 67 bp of inserted intronic sequence that may have originated due to a fork stalling and template switching/microhomology-mediated break-induced replication (FoSTeS/MMBIR) mechanism. This mechanism has not been previously implicated in HL or STL. This is also the first reported copy number variation in COL9A1 that was identified through an exome data set in an Iranian family with apparent non-syndromic HL. The present study emphasizes the importance of exome-wide copy number variation analysis in molecular diagnosis and provides supporting evidence to associate COL9A1 with autosomal recessive non-syndromic HL.     

Exome sequencing reveals a novel COL2A1 mutation implicated in multiple epiphyseal dysplasia

Mutations in the COMP, COL9A1, COL9A2, COL9A3, MATN3, and SLC26A2 genes cause approximately 70% of multiple epiphyseal dysplasia (MED) cases. The genetic changes involved in the etiology of the remaining cases are still unknown, suggesting that other genes contribute to MED development. Our goal was to identify a mutation causing an autosomal dominant form of MED in a large multigenerational family. Initially, we excluded all genes known to be associated with autosomal dominant MED by using microsatellite and SNP markers. Follow‐up with whole‐exome sequencing analysis revealed a mutation c.2032G>A (p.Gly678Arg) in the COL2A1 gene (NCBI Reference Sequence: NM_001844.4), which co‐segregated with the disease phenotype in this family, manifested by severe hip dysplasia and osteoarthritis. One of the affected family members had a double‐layered patella, which is frequently seen in patients with autosomal recessive MED caused by DTDST mutations and sporadically in the dominant form of MED caused by COL9A2 defect.     

Autosomal dominant inheritance in a recently described ZMIZ1‐related neurodevelopmental disorder: Case report of siblings and an affected parent

ZMIZ1, zinc finger MIZ‐domain containing 1, has recently been described in association with syndromic intellectual disability in which the primary phenotypic features include intellectual disability/developmental delay, seizures, hearing loss, behavioral issues, failure to thrive, and various congenital malformations. Most reported cases have been found to result from de novo mutations except for one set of three siblings in which parental testing could not be performed. With informed consent from the family, we report on a father and his two sons demonstrating autosomal dominant inheritance of a novel pathogenic ZMIZ1 variant, c.1310delC (p.Pro437ArgfsX84), causing this recently described neurodevelopmental syndrome. While they all show syndromic findings along with short stature and intellectual disability, only one child had sensorineural hearing loss. Moreover, severity of intellectual disability and eyelid ptosis were variable among the affected members. Our report demonstrates that phenotypic features of ZMIZ1‐related neurodevelopmental syndrome are variable even within the same family and that parental testing to identify a mildly affected parent is needed.     

Genotypes and estimated prevalence of phosphomannomutase 2 deficiency in Turkey differ significantly from those in Europe

Phosphomannomutase 2 deficiency (PMM2‐CDG) is an autosomal recessive congenital disorder of glycosylation, characterized by multisystem phenotypes, mostly including neurological involvement. In Turkey, due to high rates of consanguinity, many patients with autosomal recessive disorders have homozygous variants and these diseases are more common, compared to Europe. However, published reports of PMM2‐CDG from Turkey are scarce. Here, we describe clinical and molecular characteristics of PMM2‐CDG patients diagnosed in three centers in Turkey, using data obtained retrospectively from hospital records. We also analyzed an in‐house exome database of 1,313 individuals for PMM2 variants and estimated allele, carrier and disease frequencies, using the Hardy–Weinberg law. Eleven patients were identified from 10 families, displaying similar characteristics to previous publications, with the exception of the first report of epilepsia partialis continua and increased prevalence of sensorineural hearing loss. p.Val231Met was the most common variant, and was homozygous in four patients. This novel genotype results in a neurological phenotype with subclinical visceral involvement. Exome database analysis showed an estimated prevalence of 1:286,726 for PMM2‐CDG, which is much lower than expected (1:20,000 in Europe) because of the lack of predominance of the common European p.Asp141His allele, associated with a severe phenotype (allele frequency of 1:2,622 compared to 1:252 in gnomAD). These data suggest that prevalence, phenotypes and genotypes of PMM2‐CDG in Turkey differ significantly from those in Europe: Milder phenotypes may be more common, but the disease itself rarer, requiring a higher clinical suspicion for diagnosis. The association of sensorineural hearing loss with PMM2‐CDG warrants further study.     

Exome sequencing identifies homozygous variants in MBOAT7 associated with neurodevelopmental disorder

Intellectual disability (ID) is a large group of neurodevelopmental disorders characterized by a congenital limitation in intellectual functioning (reasoning, learning, and problem solving), adaptive behavior (conceptual, social, and practical skills), originated at birth and manifested before the age of 18. By whole exome sequencing of five consanguineous Pakistani families presenting hallmark features of ID, global developmental delay, aggressive and self-injurious behaviors, microcephaly, febrile seizures and facial dysmorphic features, we identified three novel homozygous missense variants (NM_024298.5: c.588G > T; p.Trp196Cys, c.736 T > C; p.Tyr246His and c.524A > C; p. Asp175Ala) and one rare homozygous in-frame deletion variant (c.758_778del;p.Glu253_Ala259del) in membrane-bound O-acyltransferase family member 7 (MBOAT7) gene previously associated with autosomal recessive neurodevelopmental disorder. The segregation of the variants was validated by Sanger sequencing in all family members. In silico homology modeling of wild-type and mutated proteins revealed substantial changes in the structure of both proteins, indicating a possible effect on function. The identification and validation of new pathogenic MBOAT7 variants in five cases of autosomal recessive ID further highlight the importance of this genes in proper brain function and development.     

A homozygous missense variant in the PLCB4 gene causes severe phenotype of auriculocondylar syndrome type 2

Auriculocondylar syndrome (ARCND) is a rare craniofacial birth defect characterized by malformations in the mandible and external ear (Question Mark Ear). Genetically, three distinct subtypes of ARCND (ARCND1, ARCND2, and ARCND3) have been identified. ARCND2 is linked to pathogenic variants in the PLCB4 gene (phospholipase C β4). PLCB4 is a key effector of the EDN1-EDNRA pathway involved in craniofacial development via the induction, migration, and maintenance of neural crest cells. ARCND2 is typically inherited in an autosomal dominant pattern, with recessive inheritance pattern being rare. In this study, we report the first homozygous missense variant (NM_000933.4: c.2050G>A: p.(Gly684Arg)) in the PLCB4 gene causing ARCND in a 3-year-old patient with a severe clinical phenotype of the syndrome. The patient presented with typical craniofacial ARCND features, in addition to intestinal transit defect, macropenis, and hearing loss. These findings further delineate the phenotypic spectrum of ARCND associated with autosomal recessive PLCB4 loss of function variants. Notably, our results provide further evidence that these variants can result in a more severe and diverse manifestations of the syndrome. Clinicians should consider the rare features of this condition for better management of patients.     

Homozygous splice site variant affecting the first von Willebrand factor A domain of COL12A1 in a patient with myopathic Ehlers-Danlos syndrome

Myopathic Ehlers-Danlos syndrome (mEDS) is a subtype of EDS that is caused by abnormalities in COL12A1. Up-to-date, 24 patients from 15 families with mEDS have been reported, with 14 families showing inheritance in an autosomal dominant manner and one family in an autosomal recessive manner. We encountered an additional patient with autosomal recessive mEDS. The patient is a 47-year-old Japanese man, born to consanguineous parents with no related features of mEDS. After birth, he presented with hypotonia, weak spontaneous movements, scoliosis, and torticollis. He had soft palms but no skin hyperextensibility or fragility. Progressive scoliosis, undescended testes, and muscular torticollis required surgery. During adulthood, he worked normally and had no physical concerns. Clinical exome analysis revealed a novel homozygous variant in COL12A1 (NM_004370.6:c.395-1G > A) at the splice acceptor site of exon 6, leading to in-frame skipping of exon 6. The patient was diagnosed with mEDS. The milder manifestations in the current patient compared with previously reported patients with mEDS might be related to the site of the variant. The variant is located in the genomic region encoding the first von Willebrand factor A domain, which affects only the long isoform of collagen XII, in contrast to the variants in previously reported mEDS patients that affected both the long and short isoforms. Further studies are needed to delineate comprehensive genotype–phenotype correlation of the disorder.     

2 new cases of pontocerebellar hypoplasia type 10 identified by whole exome sequencing in a Turkish family

Pontocerebellar hypoplasia type 10 (PCH10) is a progressive autosomal recessive neurodegenerative disorder that has been recently described in association with cleavage and polyadenylation factor I subunit 1 (CLP1) mutations. To date, all reported cases have the same homozygous missense mutation in the CLP1 gene suggesting a founder mutation. CLP1 is an RNA kinase involved in tRNA splicing and maturation. There is evidence that the mutation is associated with functionally impaired kinase activity and subsequent defective tRNA processing. Through whole exome sequencing, we identified the same mutation in an extended family of Turkish origin. Both children presented with severe psychomotor delay, progressive microcephaly, and constipation. However, intrafamilial phenotypic variability is suggested due to the variability in their brain abnormalities and clinical features.     

A Syrian patient with Steel syndrome due to compound heterozygous COL27A1 mutations with colobomata of the eye

The joint occurrence of short stature, congenital dislocation of the hip, carpal coalition, dislocation of the radial head, cavus deformity, scoliosis, and vertebral anomalies was first described in 1993 by Steel et al. (OMIM #615155) in 23 children from Puerto Rico. The condition is caused by a deficient matrix protein, collagen type XXVII alpha 1 chain, due to bi‐allelic loss of function mutations in the gene COL27A1. Outside of Puerto Rico, only four families have been described, in three of which the patients also had hearing loss. However, structural eye defects have not yet been reported in conjunction with this rare autosomal recessive syndrome. Here, we describe a 9‐year‐old girl born to nonconsanguineous Syrian parents with the characteristic features of Steel syndrome, including short stature, massive malalignment of large joints, kyphoscoliosis, hearing loss, and typical facial dysmorphism. However, she was also born with bilateral colobomata of the irides and choroido‐retinae with unilateral affection of the macula. Whole exome sequencing identified two pathogenic compound heterozygous variants in COL27A1: c.93del, p.(Phe32Leufs*71) and c.3075del, p.(Lys1026Argfs*33). There was no discernible alternative cause for the colobomata. Our findings might indicate an association of this exceptionally rare disorder caused by COL27A1 mutations with developmental defects of the eye from the anophthalmia/microphthalmia/coloboma spectrum.     

p.Arg69Trp in RNASEH2C is a founder variant in three Indian families with Aicardi–Goutières syndrome

Aicardi–Goutières syndrome is an early‐onset severe neurological disorder characterized by intracranial calcification, white matter abnormalities, hepatosplenomegaly, cerebrospinal fluid lymphocytosis, and elevated interferon‐α levels, thus mimicking congenital viral infections. It is a genetically heterogeneous condition and autosomal recessive and autosomal dominant forms with variations in seven genes known till date. Variations in RNASEH2C cause an autosomal recessive form of AGS. Here we report three Indian families with variant, c.205C>T (NM_032193.3, p.Arg69Trp) in RNASEH2C gene identified by whole‐exome sequencing and targeted molecular testing of the variant. Review of literature and our data suggest this is likely to be a founder variant in Asians and it would be a good initial variant to screen in patients with Aicardi–Goutières syndrome in Indians.     

A novel PAX1 null homozygous mutation in autosomal recessive otofaciocervical syndrome associated with severe combined immunodeficiency

Otofaciocervical syndrome (OFCS) is a rare disorder characterized by facial anomalies, cup‐shaped low‐set ears, preauricular fistulas, hearing loss, branchial defects, skeletal anomalies, and mild intellectual disability. Autosomal dominant cases are caused by deletions or point mutations of EYA1. A single family with an autosomal recessive form of OFCS and a homozygous missense mutation in PAX1 gene has been described. We report whole exome sequencing of 4 members of a consanguineous family in which 2 children, showing features of OFCS, expired from severe combined immunodeficiency (SCID). To date, the co‐occurrence of OFCS and SCID has never been reported. We found a nonsense homozygous mutation in PAX1 gene in the 2 affected children. In mice, Pax1 is required for the formation of specific skeletal structures as well as for the development of a fully functional thymus. The mouse model strongly supports the hypothesis that PAX1 depletion in our patients caused thymus aplasia responsible for SCID. This report provides evidence that bi‐allelic null PAX1 mutations may lead to a multi‐system autosomal recessive disorders, where SCID might represent the main feature.     

Expanding the phenotype spectrum associated with pathogenic variants in the COL2A1 and COL11A1 genes

We report the clinical findings of 26 individuals from 16 unrelated families carrying variants in the COL2A1 or COL11A1 genes. Using Sanger and next-generation sequencing, 11 different COL2A1 variants (seven novel), were identified in 13 families (19 affected individuals), all diagnosed with Stickler syndrome (STL) type 1. In nine families, the COL2A1 disease-causing variant arose de novo. Phenotypically, we observed myopia (95%) and retinal detachment (47%), joint hyperflexibility (92%), midface retrusion (84%), cleft palate (53%), and various degrees of hearing impairment (50%). One patient had a splenic artery aneurysm. One affected individual carrying pathogenic variant in COL2A1 showed no ocular signs including no evidence of membranous vitreous anomaly. In three families (seven affected individuals), three novel COL11A1 variants were found. The propositus with a de novo variant showed an ultrarare Marshall/STL overlap. In the second family, the only common clinical sign was postlingual progressive sensorineural hearing impairment (DFNA37). Affected individuals from the third family had typical STL2 signs. The spectrum of disease phenotypes associated with COL2A1 or COL11A1 variants continues to expand and includes typical STL and various bone dysplasias, but also nonsyndromic hearing impairment, isolated myopia with or without retinal detachment, and STL phenotype without clinically detectable ocular pathology.     

Neu–Laxova syndrome presenting prenatally with increased nuchal translucency and cystic hygroma: The utility of exome sequencing in deciphering the diagnosis

Neu–Laxova syndrome (NLS) is a lethal autosomal recessive microcephaly syndrome associated with intrauterine growth restriction (IUGR) and multiple congenital anomalies. Clinical features include central nervous system malformations, joint contractures, ichthyosis, edema, and dysmorphic facial features. Biallelic pathogenic variants in either the PHGDH or PSAT1 genes have been shown to cause NLS. Using exome sequencing, we aimed to identify the underlying genetic diagnosis in three fetuses (from one family) with prenatal skin edema, severe IUGR, micrognathia, renal anomalies, and arthrogryposis and identified a homozygous c.1A>C (p.Met1?, NM_006623.3) variant in the PHGDH gene. Loss of the translation start codon is a novel genetic mechanism for the development of NLS. Prenatal diagnosis of NLS is challenging and few reports describe the fetal pathology. Fetal neuropathologic examination revealed: delayed brain development, congenital agenesis of the corticospinal tracts, and hypoplasia of the hippocampus, cerebellum and brainstem. Each pregnancy also showed increased nuchal translucency (NT) or cystic hygroma. While NLS is rare, it may be a cause of recurrent increased NT/cystic hygroma. This finding provides further support that cystic hygroma has many different genetic causes and that exome sequencing may shed light on the underlying genetic diagnoses in this group of prenatal patients.     

Autosomal recessive otofaciocervical syndrome type 2 with novel homozygous small insertion in PAX1 gene

Otofaciocervical syndrome (OTFCS) is described as a single gene disorder of both autosomal dominant and autosomal recessive inheritance. The major clinical features of OTFCS include ear malformations (external/middle/inner ear), facial dysmorphism, shoulder girdle abnormalities, vertebral anomalies, and mild intellectual disability. The autosomal recessive form of OTFCS syndrome (OTFCS2) has been recently reported to be caused due to homozygous mutations in PAX1 gene. Here we report a third family of OTFCS2 phenotype wherein whole exome sequencing identified a novel homozygous small insertion in PAX1 as the underlying genetic cause.     

Biallelic novel missense HHAT variant causes syndromic microcephaly and cerebellar‐vermis hypoplasia

We report two siblings with microcephaly, early infantile onset seizures, and cerebellar vermis hypoplasia, in whom whole exome sequencing revealed a novel homozygous missense (c.770T>C, p.[Leu257Pro]) variant in the hedgehog acyl‐transferase gene (HHAT), encoding an enzyme required for the attachment of palmitoyl residues that are critical for multimerization and long and short range hedgehog signaling. There is a report of one family with Nivelon–Nivelon–Mabille syndrome in which HHAT was proposed as the likely candidate gene. The phenotypic overlap with the family we report herein provides further evidence implicating HHAT in cerebellar development and the pathogenesis of this rare spectrum.     

Splice-altering variant in COL11A1 as a cause of nonsyndromic hearing loss DFNA37

PurposeThe aim of this study was to determine the genetic cause of autosomal dominant nonsyndromic hearing loss segregating in a multigenerational family.MethodsClinical examination, genome-wide linkage analysis, and exome sequencing were carried out on the family.ResultsAffected individuals presented with early-onset progressive mild hearing impairment with a fairly flat, gently downsloping or U-shaped audiogram configuration. Detailed clinical examination excluded any additional symptoms. Linkage analysis detected an interval on chromosome 1p21 with a logarithm of the odds (LOD) score of 8.29: designated locus DFNA37. Exome sequencing identified a novel canonical acceptor splice-site variant c.652-2A>C in the COL11A1 gene within the DFNA37 locus. Genotyping of all 48 family members confirmed segregation of this variant with the deafness phenotype in the extended family. The c.652-2A>C variant is novel, highly conserved, and confirmed in vitro to alter RNA splicing.ConclusionWe have identified COL11A1 as the gene responsible for deafness at the DFNA37 locus. Previously, COL11A1 was solely associated with Marshall and Stickler syndromes. This study expands its phenotypic spectrum to include nonsyndromic deafness. The implications of this discovery are valuable in the clinical diagnosis, prognosis, and treatment of patients with COL11A1 pathogenic variants.