Abhijit Dixit

Mutations in CCDC39 and CCDC40 are the major cause of primary ciliary dyskinesia with axonemal disorganization and absent inner dynein arms.

Primary ciliary dyskinesia (PCD) is a genetically heterogeneous disorder caused by cilia and sperm dysmotility. About 12% of cases show perturbed 9+2 microtubule cilia structure and inner dynein arm (IDA) loss, historically termed “radial spoke defect.” We sequenced CCDC39 and CCDC40 in 54 “radial spoke defect” families, as these are the two genes identified so far to cause this defect. We discovered biallelic mutations in a remarkable 69% (37/54) of families, including identification of 25 (19 novel) mutant alleles (12 in CCDC39 and 13 in CCDC40). All the mutations were nonsense, splice, and frameshift predicting early protein truncation, which suggests this defect is caused by “null” alleles conferring complete protein loss. Most families (73%; 27/37) had homozygous mutations, including families from outbred populations. A major putative hotspot mutation was identified, CCDC40 c.248delC, as well as several other possible hotspot mutations. Together, these findings highlight the key role of CCDC39 and CCDC40 in PCD with axonemal disorganization and IDA loss, and these genes represent major candidates for genetic testing in families affected by this ciliary phenotype. We show that radial spoke structures are largely intact in these patients and propose this ciliary ultrastructural abnormality be referred to as “IDA and microtubular disorganisation defect,” rather than “radial spoke defect.”

EXOSC3 mutations in pontocerebellar hypoplasia type 1: novel mutations and genotype-phenotype correlations

BackgroundPontocerebellar hypoplasia (PCH) represents a group of neurodegenerative disorders with prenatal onset. Eight subtypes have been described thus far (PCH1-8) based on clinical and genetic features. Common characteristics include hypoplasia and atrophy of the cerebellum, variable pontine atrophy, and severe mental and motor impairments. PCH1 is distinctly characterized by the combination with degeneration of spinal motor neurons. Recently, mutations in the exosome component 3 gene (EXOSC3) have been identified in approximately half of the patients with PCH subtype 1.MethodsWe selected a cohort of 99 PCH patients (90 families) tested negative for mutations in the TSEN genes, RARS2, VRK1 and CASK. Patients in this cohort were referred with a tentative diagnose PCH type 1, 2, 4, 7 or unclassified PCH. Genetic analysis of the EXOSC3 gene was performed using Sanger sequencing. Clinical data, MR images and autopsy reports of patients positive for EXOSC3 mutations were analyzed.ResultsEXOSC3 mutations were found in twelve families with PCH subtype 1, and were not found in patients with other PCH subtypes. Identified mutations included a large deletion, nonsense and missense mutations. Examination of clinical data reveals a prolonged disease course in patients with a homozygous p.D132A mutation. MRI shows variable pontine hypoplasia in EXOSC3 mediated PCH, where the pons is largely preserved in patients with a homozygous p.D132A mutation, but attenuated in patients with other mutations. Additionally, bilateral cerebellar cysts were found in patients compound heterozygous for a p.D132A mutation and a nonsense allele.ConclusionsEXOSC3 mediated PCH shows clear genotype-phenotype correlations. A homozygous p.D132A mutation leads to PCH with possible survival into early puberty, and preservation of the pons. Compound heterozygosity for a p.D132A mutation and a nonsense or p.Y109N allele, a homozygous p.G31A mutation or a p.G135E mutation causes a more rapidly progressive course leading to death in infancy and attenuation of the ventral pons.Our findings imply a clear correlation between genetic mutation and clinical outcome in EXOSC3 mediated PCH, including variable involvement of the pons.

A Restricted Repertoire of De Novo Mutations in ITPR1 Cause Gillespie Syndrome with Evidence for Dominant-Negative Effect

Gillespie syndrome (GS) is characterized by bilateral iris hypoplasia, congenital hypotonia, non-progressive ataxia, and progressive cerebellar atrophy. Trio-based exome sequencing identified de novo mutations in ITPR1 in three unrelated individuals with GS recruited to the Deciphering Developmental Disorders study. Whole-exome or targeted sequence analysis identified plausible disease-causing ITPR1 mutations in 10/10 additional GS-affected individuals. These ultra-rare protein-altering variants affected only three residues in ITPR1: Glu2094 missense (one de novo, one co-segregating), Gly2539 missense (five de novo, one inheritance uncertain), and Lys2596 in-frame deletion (four de novo). No clinical or radiological differences were evident between individuals with different mutations. ITPR1 encodes an inositol 1,4,5-triphosphate-responsive calcium channel. The homo-tetrameric structure has been solved by cryoelectron microscopy. Using estimations of the degree of structural change induced by known recessive- and dominant-negative mutations in other disease-associated multimeric channels, we developed a generalizable computational approach to indicate the likely mutational mechanism. This analysis supports a dominant-negative mechanism for GS variants in ITPR1. In GS-derived lymphoblastoid cell lines (LCLs), the proportion of ITPR1-positive cells using immunofluorescence was significantly higher in mutant than control LCLs, consistent with an abnormality of nuclear calcium signaling feedback control. Super-resolution imaging supports the existence of an ITPR1-lined nucleoplasmic reticulum. Mice with Itpr1 heterozygous null mutations showed no major iris defects. Purkinje cells of the cerebellum appear to be the most sensitive to impaired ITPR1 function in humans. Iris hypoplasia is likely to result from either complete loss of ITPR1 activity or structure-specific disruption of multimeric interactions.

The Koolen-de Vries syndrome: A phenotypic comparison of patients with a 17q21.31 microdeletion versus a KANSL1 sequence variant

The Koolen-de Vries syndrome (KdVS; OMIM #610443), also known as the 17q21.31 microdeletion syndrome, is a clinically heterogeneous disorder characterised by (neonatal) hypotonia, developmental delay, moderate intellectual disability, and characteristic facial dysmorphism. Expressive language development is particularly impaired compared with receptive language or motor skills. Other frequently reported features include social and friendly behaviour, epilepsy, musculoskeletal anomalies, congenital heart defects, urogenital malformations, and ectodermal anomalies. The syndrome is caused by a truncating variant in the KAT8 regulatory NSL complex unit 1 (KANSL1) gene or by a 17q21.31 microdeletion encompassing KANSL1. Herein we describe a novel cohort of 45 individuals with KdVS of whom 33 have a 17q21.31 microdeletion and 12 a single-nucleotide variant (SNV) in KANSL1 (19 males, 26 females; age range 7 months to 50 years). We provide guidance about the potential pitfalls in the laboratory testing and emphasise the challenges of KANSL1 variant calling and DNA copy number analysis in the complex 17q21.31 region. Moreover, we present detailed phenotypic information, including neuropsychological features, that contribute to the broad phenotypic spectrum of the syndrome. Comparison of the phenotype of both the microdeletion and SNV patients does not show differences of clinical importance, stressing that haploinsufficiency of KANSL1 is sufficient to cause the full KdVS phenotype.

7q11.23 Microduplication: a recognizable phenotype

Dixit A, McKee S, Mansour S, Mehta SG, Tanteles GA, Anastasiadou V, Patsalis PC, Martin K, McCullough S, Suri M, Sarkar A. 7q11.23 Microduplication: a recognizable phenotype.

17q12 Microdeletion Syndrome: Three Patients Illustrating the Phenotypic Spectrum

Deletions of 17q12 are associated with renal cysts and maturity onset diabetes of the young, and have also been identified in women with reproductive tract anomalies due to Mullerian aplasia. Although initially identified in patients with normal cognitive ability, some patients with this recurrent microdeletion syndrome have learning problems. We identified a 17q12 microdeletion in three patients with renal cystic disease by array comparative genomic hybridization and the phenotypic spectrum of the 17q12 microdeletion syndrome is illustrated by the description of these patients. Of two patients who are old enough to be assessed, one has significant speech delay, autism spectrum disorder, and mild learning difficulty, while the other patient has only mild speech delay. This highlights the variability of cognitive involvement in this condition. The third patient presented with Alagille syndrome‐like features in the neonatal period. All three patients had transient hypercalcemia in the neonatal period, a finding that has not previously been described in this condition. Moreover, two patients have mild or no dysmorphism, while one displays striking facial dysmorphism in addition to minor congenital anomalies. We suggest that while patients with 17q12 microdeletion syndrome can present with type 2 diabetes or renal cysts without any dysmorphic features, a subgroup may have dysmorphic features or present with neonatal cholestasis. Transient neonatal hypercalcemia may be a feature of this microdeletion syndrome.

Monozygotic twins discordant for trisomy 13: counselling and management issues.

The diagnosis and management of a heterokaryotypic monochorionic pregnancy, in which one of twins had trisomy 13, is presented. Monozygosity and discordant karyotypes were confirmed by amniocentesis of both the sacs. Radiofrequency ablation of the trisomic twin was successfully performed at 18-weeks gestation and the pregnancy ended at term with the birth of a healthy girl who remains well on follow-up at 12 months of age. We reiterate the importance of early amniocentesis of both the sacs in the presence of discordant fetal abnormalities and consideration of selective fetal termination to optimise the outcome of heterokaryotypic monochorionic twin pregnancies.

Mild phenotype in a patient with a de-novo 2.9-Mb interstitial deletion at 13q12.11.

Introduction Interstitial deletions of 13q12 are extremely rare. Increasing clinical application of relatively new technologies such as microarray-comparative genomic hybridization [microarray-CGH)] is proving to be a powerful tool in identifying submicroscopic chromosome deletions and duplications that can manifest with great phenotypic variability. We describe a 16-year-old boy with a de-novo 2.9-Mb interstitial deletion at 13q12.11, with previously unreported breakpoints and a relatively mild phenotype with a degree of facial dysmorphism, scaphocephaly, torticollis and near normal development and intellect.

ACTB Loss-of-Function Mutations Result in a Pleiotropic Developmental Disorder

ACTB encodes β-actin, an abundant cytoskeletal housekeeping protein. In humans, postulated gain-of-function missense mutations cause Baraitser-Winter syndrome (BRWS), characterized by intellectual disability, cortical malformations, coloboma, sensorineural deafness, and typical facial features. To date, the consequences of loss-of-function ACTB mutations have not been proven conclusively. We describe heterozygous ACTB deletions and nonsense and frameshift mutations in 33 individuals with developmental delay, apparent intellectual disability, increased frequency of internal organ malformations (including those of the heart and the renal tract), growth retardation, and a recognizable facial gestalt (interrupted wavy eyebrows, dense eyelashes, wide nose, wide mouth, and a prominent chin) that is distinct from characteristics of individuals with BRWS. Strikingly, this spectrum overlaps with that of several chromatin-remodeling developmental disorders. In wild-type mouse embryos, β-actin expression was prominent in the kidney, heart, and brain. ACTB mRNA expression levels in lymphoblastic lines and fibroblasts derived from affected individuals were decreased in comparison to those in control cells. Fibroblasts derived from an affected individual and ACTB siRNA knockdown in wild-type fibroblasts showed altered cell shape and migration, consistent with known roles of cytoplasmic β-actin. We also demonstrate that ACTB haploinsufficiency leads to reduced cell proliferation, altered expression of cell-cycle genes, and decreased amounts of nuclear, but not cytoplasmic, β-actin. In conclusion, we show that heterozygous loss-of-function ACTB mutations cause a distinct pleiotropic malformation syndrome with intellectual disability. Our biological studies suggest that a critically reduced amount of this protein alters cell shape, migration, proliferation, and gene expression to the detriment of brain, heart, and kidney development.

Two somali half-siblings with CHST3-related chondrodysplasia illustrating the phenotypic spectrum and intrafamilial variability

Deficiency of carbohydrate sulfotransferase 3 (CHST3; also known as chondroitin‐6‐sulfotranferase) has been associated with a phenotype of severe chondrodysplasia and progressive spinal involvement. Recent reports indicate that affected individuals initially present with neonatal multiple joint dislocations. We describe a 14‐year‐old Somali patient and her 3‐year‐old maternal half‐brother with novel homozygous CHST3 mutations. The proband presented at the age 5½ years with short stature and genua valga. Her clinical course was characterized by rapid progression of spinal deformities and large joint contractures. Her half‐brother presented at birth with bilateral knee dislocation and talipes equinovarus. This report of a Somali family with CHST3‐related chondrodysplasia illustrates the intrafamilial variability in phenotypic expression of this rare disorder.