Translocation Is the Likely Driver of a Syndrome with Ambiguous Genitalia, Facial Dysmorphism, Intellectual Disability, and Speech Delay

COLD SPRING HARBOR Molecular Case Studies | RESEARCH REPORT A t(5;16) translocation is the likely driver of a syndrome with ambiguous genitalia, facial dysmorphism, intellectual disability, and speech delay Ays¸egül Ozantürk,1,2,3 Erica E. Davis,3 Aniko Sabo,4 Marjan M. Weiss,5 Donna Muzny,4 Shannon Dugan-Perez,4 Erik A. Sistermans,5 Richard A. Gibbs,4 Köksal R. Özgül,1 Dilek Yalnızoglu,6 Esra Serdaroglu,6 Ali Dursun,1 and Nicholas Katsanis3 1Department of Pediatrics, Metabolism Unit, Hacettepe University, Ankara 06410, Turkey; 2Department of Biology, Molecular Biology Section, Hacettepe University, Ankara 06800, Turkey; 3Center for Human Disease Modeling, Duke University, Durham, North Carolina 27701, USA; 4Human Genome Sequencing Center, Baylor College of Medicine, Houston, Texas 77030, USA; 5Department of Clinical Genetics, VU University Medical Center (Amsterdam), NL-1081 HV Amsterdam, The Netherlands; 6Department of Pediatrics, Pediatric Neurology Unit, Hacettepe University, Ankara 06410, Turkey Abstract Genetic studies grounded on monogenic paradigms have accelerated both gene discovery and molecular diagnosis. At the same time, complex genomic rearrangements are Corresponding authors: also appreciated as potent drivers of disease pathology. Here, we report two male siblings [email protected]; [email protected] with a dysmorphic face, ambiguous genitalia, intellectual disability, and speech delay. Through quad-based whole-exome sequencing and concomitant molecular cytogenetic testing, we identified two copy-number variants (CNVs) in both affected individuals © 2016 Ozantürk et al. This article likely arising from a balanced translocation: a 13.5-Mb duplication on Chromosome 16 is distributed under the terms of → → the Creative Commons (16q23.1 16qter) and a 7.7-Mb deletion on Chromosome 5 (5p15.31 5pter), as well as Attribution-NonCommercial a hemizygous missense variant in CXorf36 (also known as DIA1R). The 5p terminal deletion License, which permits reuse and has been associated previously with speech delay, whereas craniofacial dysmorphia redistribution, except for and genital/urinary anomalies have been reported in patients with a terminal duplication commercial purposes, provided of 16q. However, dosage changes in either genomic region alone could not account that the original author and for the overall clinical presentation in our family; functional testing of CXorf36 in zebrafish source are credited. did not induce defects in neurogenesis or the craniofacial skeleton. Notably, literature and database analysis revealed a similar dosage disruption in two siblings with extensive Ontology terms: cat cry; central phenotypic overlap with our patients. Taken together, our data suggest that dosage hypotonia; clubbing of toes; perturbation of genes within the two chromosomal regions likely drives the syndromic delayed gross motor manifestations of our patients and highlight how multiple genetic lesions can contribute to development; down-sloping shoulders; enlarged proximal complex clinical pathologies. interphalangeal joints; high, narrow palate; intellectual disability, profound; micropenis; [Supplemental material is available for this article.] moderately short stature; penile hypospadias; pes planus; prominent forehead; thin upper INTRODUCTION lip vermilion; wide nasal bridge Whole-exome sequencing (WES) has become a commonly used tool for both gene discov- Published by Cold Spring Harbor ery in the research setting and accelerated diagnostics in the clinical setting. Reports from Laboratory Press ever-increasing patient cohorts have indicated that WES can deliver a molecular diagnosis doi: 10.1101/mcs.a000703 ∼25% of the time (Yang et al. 2013; Beaulieu et al. 2014). In parallel, the combinatorial Cite this article as Ozantürk et al. 2016 Cold Spring Harb Mol Case Stud 2: a000703 1 of 14 COLD SPRING HARBOR Molecular Case Studies Unbalanced t(5;16) translocation in a new syndrome application of oligo-based comparative genomic hybridization arrays and computational analyses that infer chromosomal losses/gains from nonuniform read depth have also high- lighted an important contribution of copy-number variants (CNVs) to rare genetic disorders (Solinas-Toldo et al. 1997; Snijders et al. 2001; Schaaf et al. 2011). Despite these advances, a significant fraction of patients tested by these methods yields no overt candidates, even in cases in which the inheritance of the phenotype within a family suggests a strong underlying genetic component. There are many reasons for this, including the persistent difficulties in interpreting rare variation, the low quality or coverage of some regions of the genome, the poor detection of some CNVs, and the contribution of mutations in noncoding regulatory elements. In addition, examples have emerged in which a single- gene–single-syndrome paradigm is not sufficient to explain the inheritance of clinical phe- notypes. In some cases, the clinical presentation represents two overlapping disorders, thus confusing locus assignment (Ng et al. 2010; Roach et al. 2010; Yang et al. 2013). In other cases, alleles can interact with each other at discrete loci to modulate penetrance and/or ex- pressivity. These can represent genetic interactions of either single-nucleotide variants (SNVs) (Davis and Katsanis 2012), SNVs with CNVs (Lemmers et al. 2012; Lindstrand et al. 2014), or, in principle, CNVs with CNVs (Girirajan et al. 2012). Here, we report the combinatorial results of quad-based WES and molecular cytogenetic analysis of Turkish male siblings with a constellation of neurocognitive, craniofacial, and go- nadal malformations that do not conform readily to an established syndrome or known molecular lesion. The aggregate analysis for SNVs and CNVs identified two dosage perturbations in both affected individuals—a terminal duplication on Chromosome 16q and a terminal deletion on Chromosome 5p—as well as a maternally inherited missense variant (p.Ser105Pro) in CXorf36. Functional studies in zebrafish embryos indicated that perturbation of CXorf36 is unlikely to contribute to the patients’ dysmorphic features or cause neurodevelopmental abnormalities, whereas molecular cytogenetic analysis indicated that no gene was disrupted in either CNV boundary. In contrast, retrospective analysis of reported cases led to the identification of a family of Saudi Arabian descent who shared clinical features with our patients and had likewise overlapping 5p loss and 16q gain (Hellani et al. 2010). Some clinical features can potentially be explained by dosage defects on either chromosomal location. However, the remaining defects in our patients, which are shared by the previously published family, argue that concomitant haploinsufficiency on 5p and increased gene dosage across 16q constitute the most parsimonious driver hypothesis for this syndrome. RESULTS Clinical Characterization of a Syndromic Disorder in a Turkish Pedigree We consulted for a nonconsanguineous Turkish family with two male siblings with the prima- ry features of motor delay accompanied by intellectual disability and ambiguous genitalia (Table 1; Fig. 1A) at Hacettepe University Hospital. The parents reported no family history of inherited disease, except for a deceased female child who was affected with hypotonia of unknown etiology. We evaluated the oldest affected child (M-11-1496) when he was 11 yr, 4 mo old. Physical examination showed a weight of 26.5 kg (<5th percentile), height of 124 cm (<5th percentile), and head circumference of 51 cm. He had a weak cry (cat-like), stridor, and hypotonia at birth; his stridor recovered when he was 18 mo of age. He had delayed developmental milestones: He was able to hold his head up and sit by himself at 4 yr and he walked at 9.5 yr. First, we noted focal neurological deficits and stereotypic movements. He did not respond to his name nor make eye contact, and he was unable to speak. His gait is wide because of pes planus, and he displayed minimal coordination. Second, he has a micropenis, hypospadias, and undescended testes. Moreover, we noted distinct craniofacial and Ozantürk et al. 2016 Cold Spring Harb Mol Case Stud 2: a000703 2 of 14 COLD SPRING HARBOR Molecular Case Studies Unbalanced t(5;16) translocation in a new syndrome Table 1. Phenotypic comparison between individuals with 16q23 dup and 5p15 del Present report Hellani et al. 2010 M-11-1496 M-11-1497 Patient 1 Patient 2 16q23.1 → ter 16q23.1 → ter 16q23.3→ter 16q23.3→ter Phenotype 5p15.31 → ter 5p15.31 → ter 5p15.32→ter 5p15.32→ter Hypotonia ++++ HP:0001319 Congenital heart disease −−++ HP:0001635 Umbilical hernia −−++ HP:0001537 Pectus excavatum −−++ HP:0000767 Genital hypoplasia ++++ HP:0003241 Short stature ++++ HP:0004322 Hip rotation −−++ HP:0001374 Metatarsus adductus −−++ HP:0001840 Clinodactyly −−++ HP:0030084 Motor developmental delay ++++ HP:0001270 Intellectual disability ++++ HP:0001249 Brachycephaly −−++ HP:0000248 Prominent forehead ++++ HP:0011220 Broad nasal bridge ++++ HP:0000431 Low-set ears ++++ HP:0000369 Arched palate ++++ HP:0000218 Thin upper lip ++++ HP:0000177 Esotropia NR NR + + HP:0000565 Speech delay ++++ HP:0000750 Stridor + + NR NR HP:0010307 Scaphocephaly + + NR NR HP:0000268 Stereotypic movements + + NR NR HP:000733 Continued Ozantürk et al. 2016 Cold Spring Harb Mol Case Stud 2: a000703 3 of 14 COLD SPRING HARBOR Molecular Case Studies Unbalanced t(5;16) translocation in a new syndrome Table 1. Continued Present report Hellani et al. 2010 M-11-1496 M-11-1497 Patient 1 Patient

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