Ranad Shaheen

Novel CENPJ mutation causes Seckel syndrome

Background Primordial dwarfism (PD) is an extremely rare, clinicallyheterogeneous condition characterised by profound prenatal and postnatal growth restriction among other manifestations that are helpful in the clinical classification. Recently, mutation of PCNT was reported in the context of two overlapping forms of PD: Seckel syndrome and Majewskiosteodysplastic primordial dwarfism type II (MOPDII). Aim To clinically and molecularly characterise a consanguineous family with Seckel syndrome. Methods Clinical evaluation, linkage analysis, homozygosity mapping and mutation analysis. Results Unexpectedly, linkage analysis led to the identification of a novel splice-site mutation in CENPJ that segregates with the phenotype in this family. Conclusion This report establishes for the first time that mutation of CENPJ can lead to Seckel syndrome and calls for further investigation of the role played by other microcephaly related genes in the pathogenesis of PD.

Recessive Mutations in DOCK6, Encoding the Guanidine Nucleotide Exchange Factor DOCK6, Lead to Abnormal Actin Cytoskeleton Organization and Adams-Oliver Syndrome

Adams-Oliver syndrome (AOS) is defined by the combination of aplasia cutis congenita (ACC) and terminal transverse limb defects (TTLD). It is usually inherited as an autosomal-dominant trait, but autosomal-recessive inheritance has also been documented. In an individual with autosomal-recessive AOS, we combined autozygome analysis with exome sequencing to identify a homozygous truncating mutation in dedicator of cytokinesis 6 gene (DOCK6) which encodes an atypical guanidine exchange factor (GEF) known to activate two members of the Rho GTPase family: Cdc42 and Rac1. Another homozygous truncating mutation was identified upon targeted sequencing of DOCK6 in an unrelated individual with AOS. Consistent with the established role of Cdc42 and Rac1 in the organization of the actin cytoskeleton, we demonstrate a cellular phenotype typical of a defective actin cytoskeleton in patient cells. These findings, combined with a Dock6 expression profile that is consistent with an AOS phenotype as well as the very recent demonstration of dominant mutations of ARHGAP31 in AOS, establish Cdc42 and Rac1 as key molecules in the pathogenesis of AOS and suggest that other regulators of these Rho GTPase proteins might be good candidates in the quest to define the genetic spectrum of this genetically heterogeneous condition.

Study of autosomal recessive osteogenesis imperfecta in Arabia reveals a novel locus defined by TMEM38B mutation

Background Osteogenesis imperfecta (OI) is an hereditary bone disease in which increased bone fragility leads to frequent fractures and other complications, usually in an autosomal dominant fashion. An expanding list of genes that encode proteins related to collagen metabolism are now recognised as important causes of autosomal recessive (AR) OI. Our aim was to study the contribution of known genes to AR OI in order to identify novel loci in mutation-negative cases. Methods We enrolled multiplex consanguineous families and simplex cases (also consanguineous) in which mutations in COL1A1 and COL1A2 had been excluded. We used autozygome guided mutation analysis of AR OI (AR OI) genes followed by exome sequencing when such analysis failed to identify the causative mutation. Results Two simplex and 11 multiplex families were enrolled, encompassing 27 cases. In three multiplex families, autozygosity and linkage analysis revealed a novel recessive OI locus on chromosome 9q31.1-31.3, and a novel truncating deletion of exon 4 of TMEM38B was identified within that interval. In addition, gonadal or gonadal/somatic mosaic mutations in COL1A1 or COL1A2 and homozygous mutations in recently described AR OI genes were identified in all remaining families. Conclusions TMEM38B is a novel candidate gene for AR OI. Future studies are needed to explore fully the contribution of this gene to AR OI in other populations.

Identification of KLHL41 Mutations Implicates BTB-Kelch-Mediated Ubiquitination as an Alternate Pathway to Myofibrillar Disruption in Nemaline Myopathy

Nemaline myopathy (NM) is a rare congenital muscle disorder primarily affecting skeletal muscles that results in neonatal death in severe cases as a result of associated respiratory insufficiency. NM is thought to be a disease of sarcomeric thin filaments as six of eight known genes whose mutation can cause NM encode components of that structure, however, recent discoveries of mutations in non-thin filament genes has called this model in question. We performed whole-exome sequencing and have identified recessive small deletions and missense changes in the Kelch-like family member 41 gene (KLHL41) in four individuals from unrelated NM families. Sanger sequencing of 116 unrelated individuals with NM identified compound heterozygous changes in KLHL41 in a fifth family. Mutations in KLHL41 showed a clear phenotype-genotype correlation: Frameshift mutations resulted in severe phenotypes with neonatal death, whereas missense changes resulted in impaired motor function with survival into late childhood and/or early adulthood. Functional studies in zebrafish showed that loss of Klhl41 results in highly diminished motor function and myofibrillar disorganization, with nemaline body formation, the pathological hallmark of NM. These studies expand the genetic heterogeneity of NM and implicate a critical role of BTB-Kelch family members in maintenance of sarcomeric integrity in NM.

Genomic analysis of primordial dwarfism reveals novel disease genes

Primordial dwarfism (PD) is a disease in which severely impaired fetal growth persists throughout postnatal development and results in stunted adult size. The condition is highly heterogeneous clinically, but the use of certain phenotypic aspects such as head circumference and facial appearance has proven helpful in defining clinical subgroups. In this study, we present the results of clinical and genomic characterization of 16 new patients in whom a broad definition of PD was used (e.g., 3M syndrome was included). We report a novel PD syndrome with distinct facies in two unrelated patients, each with a different homozygous truncating mutation in CRIPT. Our analysis also reveals, in addition to mutations in known PD disease genes, the first instance of biallelic truncating BRCA2 mutation causing PD with normal bone marrow analysis. In addition, we have identified a novel locus for Seckel syndrome based on a consanguineous multiplex family and identified a homozygous truncating mutation in DNA2 as the likely cause. An additional novel PD disease candidate gene XRCC4 was identified by autozygome/exome analysis, and the knockout mouse phenotype is highly compatible with PD. Thus, we add a number of novel genes to the growing list of PD-linked genes, including one which we show to be linked to a novel PD syndrome with a distinct facial appearance. PD is extremely heterogeneous genetically and clinically, and genomic tools are often required to reach a molecular diagnosis.

FREM1 Mutations Cause Bifid Nose, Renal Agenesis, and Anorectal Malformations Syndrome

An autosomal-recessive syndrome of bifid nose and anorectal and renal anomalies (BNAR) was previously reported in a consanguineous Egyptian sibship. Here, we report the results of linkage analysis, on this family and on two other families with a similar phenotype, which identified a shared region of homozygosity on chromosome 9p22.2-p23. Candidate-gene analysis revealed homozygous frameshift and missense mutations in FREM1, which encodes an extracellular matrix component of basement membranes. In situ hybridization experiments demonstrated gene expression of Frem1 in the midline of E11.5 mouse embryos, in agreement with the observed cleft nose phenotype of our patients. FREM1 is part of a ternary complex that includes FRAS1 and FREM2, and mutations of the latter two genes have been reported to cause Fraser syndrome in mice and humans. The phenotypic variability previously reported for different Frem1 mouse mutants suggests that the apparently distinct phenotype of BNAR in humans may represent a previously unrecognized variant of Fraser syndrome.

Mutations in EOGT Confirm the Genetic Heterogeneity of Autosomal-Recessive Adams-Oliver Syndrome

Adams-Oliver syndrome (AOS) is a rare, autosomal-dominant or -recessive disorder characterized primarily by aplasia cutis congenita and terminal transverse limb defects. Recently, we demonstrated that homozygous mutations in DOCK6 cause an autosomal-recessive form of AOS. In this study, we sought to determine the contribution of DOCK6 mutations to the etiology of AOS in several consanguineous families. In two of the five families studied, we identified two homozygous truncating mutations (a splice-site mutation and a frameshift duplication). DOCK6 sequencing revealed no mutation in the remaining three families, consistent with their autozygosity mapping and linkage-analysis results, which revealed a single candidate locus in 3p14.1 on three different haplotype backgrounds in the three families. Indeed, exome sequencing in one family revealed one missense mutation in EOGT (C3orf64), and subsequent targeted sequencing of this gene revealed a homozygous missense mutation and a homozygous frameshift deletion mutation in the other two families. EOGT encodes EGF-domain-specific O-linked N-acetylglucosamine (O-GlcNAc) transferase, which is involved in the O-GlcNAcylation (attachment of O-GlcNAc to serine and threonine residues) of a subset of extracellular EGF-domain-containing proteins. It has a documented role in epithelial-cell-matrix interactions in Drosophila, in which deficiency of its ortholog causes wing blistering. Our findings highlight a developmental role of O-GlcNAcylation in humans and expand the genetic heterogeneity of autosomal-recessive AOS.

Clinical and molecular characterisation of Bardet–Biedl syndrome in consanguineous populations: the power of homozygosity mapping

Bardet–Biedl syndrome (BBS) is a ciliopathy with pleiotropic effect that manifests primarily as renal insufficiency, polydactyly, retinal dystrophy and obesity. The current phenotype–genotype correlation is insufficient to predict the likely causative mutation that makes sequencing of all 14 BBS genes an often necessary but highly complicated way to identify the underlying genetic defect in affected patients. In this study, homozygosity mapping is shown as a robust approach that is highly suited for genetically heterogeneous autosomal recessive disorders in populations in which consanguinity is prevalent. This approach allowed us to quickly identify seven novel mutations in seven families with BBS. Some of these mutations would have been missed by unguided routine sequencing, which suggests that missed mutations in known BBS genes could be more common than previously thought. This study, the largest to date on Saudi BBS families, also revealed interesting phenotypic aspects of BBS, including the first report of non-syndromic retinitis pigmentosa as a novel BBS phenotype.

A TCTN2 mutation defines a novel Meckel Gruber syndrome locus

Meckel Gruber syndrome (MKS) is an autosomal recessive multisystem disorder that represents a severe form of ciliopathy in humans and is characterized by significant genetic heterogeneity. In this article, we describe the identification of a novel MKS locus MKS8 that we map to TCTN2, in a multiplex consanguineous family. TCTN2 is a paralog of the recently identified Tectonic 1, which has been shown to modulate sonic hedgehog signaling. Expression analysis at different developmental stages of the murine ortholog revealed a spatial and temporal pattern consistent with the MKS phenotype observed in our patient. The exclusion of this and the other seven MKS genes in our collection of consanguineous Arab MKS families confirms the existence of two additional MKS loci. Hum Mutat 32:1–6, 2011.

Mutations in FKBP10 cause both Bruck syndrome and isolated osteogenesis imperfecta in humans.

Bruck syndrome (BS) is an autosomal recessive syndromic form of osteogenesis imperfecta (OI) that is characterized by the additional presence of pterygium formation. We have recently shown that FKBP10 previously reported as a novel autosomal recessive OI gene also defines a novel Bruck syndrome locus (BKS3). In this manuscript, we extend our analysis to describe a mutation previously described in isolated OI patients and show that it results in BS phenotype in a Saudi family. More interestingly, we describe a novel FKBP10 mutation that results in isolated OI as well as BS phenotype in the same family. These results, combined with recently published work, confirm that FKBP10 is a bonafide BS locus and lay the foundation for future research into modifiers that underlie the phenotypic heterogeneity of FKBP10 mutations.