Whole exome sequencing identified a novel truncation mutation in the NHS gene associated with Nance-Horan syndrome

Nance-Horan syndrome (NHS) is a rare X-linked genetic disorder. According to the latest statistics of the National Organization for Rare Disorders shown that fewer than 50 families have been described in the medical literatures, and the exact incidence of the disorder remains unknown (https://​rarediseases.​org/​rare-diseases/​nance-horan-syndrome/​). NHS is caused by a mutation in the NHS gene on chromosome Xp22. NHS is usually fully expressed in males only, with the affected males characterized by congenital cataracts and frequent microcornea, dental anomalies, and dysmorphic features [1, 2]. Approximately 20 to 30% of affected males may have varying levels of mental retardation [3]. Autism spectrum disorder symptoms have also been observed in a few patients [4]. Carrier females may show posterior Y-sutural cataracts with small corneas and slightly reduced vision only, they do not usually develop intellectual impairment [5, 6]. Since the symptoms of isolated microphthalmia with cataract; cataract-microcornea syndrome; X-linked congenital cataract and X-linked congenital cataract with microcornea are similar to those of NHS, making a definitive diagnosis without genetic information is difficult. Thus, to determine the potential disease-causing gene, we performed whole exome sequencing (WES) analysis in a proband.

In this study, we revealed a novel truncation mutation (c.C4449G, p.Tyr1483Ter) in the NHS gene, which might associated with NHS in a three-generation Chinese family. The whole coding sequence of NHS gene is 4893 bp, and in our study the novel truncation occurred in c.4449 site, and the affected individuals in the family presented with relatively mild abnormality. Although the male affected individuals had the same variation, the phenotypic heterogeneity existed in the NHS family (Table 1). This implied that the underlying mechanism of clinical heterogeneity was still needed to be investigated.

NHS manifestations appear in multiple organ systems especially the features of facial, teeth, skeleton, heart and neurologic system features, and even encompass behavior. And heterozygous females commonly had posterior Y-sutural cataracts. In this study we had no chance to perform the professional ophthalmologic examination for the carrier females, since they do not think they have vision impairment, and their lens opacity and vision was not quite clear. However, X-linked cataract merely involves the eyes. Heterozygous females had posterior stellate or suture cataracts, or a combination of the two, with normal vision or a slight reduction (Table 2). However, the facial dysmorphology and dental abnormalities might be subtle and easily missed in NHS patients in whom congenital cataracts are the primary clinical concern. Additionally, there was not enough evidence to identify the two disorders that might be allelic [9].

The NHS gene is alternatively spliced and composed of 10 coding exons, which encode a protein containing 4 conserved nuclear localization signals, The encoded protein can regulate actin remodeling and cell morphology. Expression studies in mice showed that the NHS gene plays critical roles in regulating eye, tooth, brain, and craniofacial development. Additionally, the complex pattern of temporally and spatially regulated expression of NHS was consistent with the pleiotropic features of NHS [3].

Previous studies illustrated that variations in the NHS gene could induce X-linked cataracts and NHS [10‐12]. However, the identical association between the genotype and phenotype was unclear. One study predicted that a lack of functional NHS protein might cause NHS, whereas aberrant transcription of the NHS gene might lead to a milder X-linked cataract phenotype [10]. To determine whether NHS and X-linked cataract were the two distinguishing phenotypes of NHS gene mutations, we reviewed the pathogenic/likely pathogenic mutations, which were recorded in the ClinVar and Cat-Map databases [13]. Here, more than 50 mutations of the NHS gene accounted for pathogenic clinical conditions. Variants including missense, frame-shift (InDel) and stop-gain mutations were identified as contributors to NHS; and a 4.8 kb deletion and a 500 kb triplication were associated with X-linked cataract 40 (Additional file 4); Considering the genotypes and phenotypes of NHS and X-linked cataracts, we found that NHS was mostly resulted from the point mutation and micro InDel [14‐16], while the X-linked cataract was caused by large-scale recombination of the NHS gene, which was predicted to result in altered transcriptional regulation of the NHS gene. However, this difference could not be used to define NHS and X-linked cataract [9], since a microdeletion of 170,6 kb at Xp22.13 was detected in an Italian boy with NHS syndrome [15]. The latest cytogenetic and molecular analyses in two patients, a mother and daughter, who presented with NHS, demonstrated a 46, X, t(X;1) (p22.13;q22) karyotype in each patient. No copy-number genomic imbalances were detected. The mother had a preferential inactivation of the normal X chromosome and no mRNA isoform of NHS was detected. This study implied that NHS could be due to the disruption of NHS gene expression [12].

NHS-affected individuals have mutations in the NHS gene that typically result in premature truncation of the protein (Additional file 4). The mutant proteins failed to localize to the cellular periphery in epithelial cells and instead were found in the cytoplasm. The mislocalization of the mutant NHS-A protein is expected to adversely affect cell-cell junctions in epithelial cells such as the lens epithelium, which may explain the cataractogenesis in NHS patients [10, 17, 18], and NHS proteins localized to the cellular periphery. In the developing of mammalian lens, researchers discovered continuous expression of NHS that became restricted to the lens epithelium in prenatal and postnatal lenses; this finding suggested that disturbances in intercellular contacts were the underlying mechanisms of cataractogenesis in NHS [19]. Additionally, the NHS gene appears to have multiple isoforms as a result of alternative transcription. The N-terminus of isoforms NHS-A and NHS-1A, which are implicated in the pathogenesis of NHS, have a functional WAVE homology domain that interacts with the Abi protein family, haematopoietic stem/progenitor cell protein 300 (HSPC300), Nap1 and Sra1. NHS knockdown resulted in the disruption of the actin cytoskeleton, and led to a striking increase in cell spreading. Conversely, ectopic over expression of NHS inhibited lamellipod formation. The NHS gene was demonstrated to be a novel regulator of actin remodeling and cell morphology, and may orchestrate actin regulatory protein function in response to signaling events during development [20]. Therefore, we speculate that X-linked cataract might represent differences in the expression of NHS.

In this study, we found a novel truncation mutation in the NHS gene, which was predicted to be likely associated with NHS. Moreover, we reviewed the characteristics of the NHS gene mutations in NHS and X-linked cataract, this review illustrated that NHS mutations in the NHS-affected individuals typically result in premature truncation of the protein or disruption of NHS gene expression; and the X-linked cataract might due to the protein expression aberration. Additionally, NHS has significantly clinical heterogeneity, and the manifestations of mild NHS could be similar to those of X-linked cataract, which suggested that the combination of genome detection and the manifestations could improve the clinical diagnosis. And, the novel mutation in the NHS gene might highlight the understanding of the causative mutations of Nance-Horan syndrome.