A novel NHS mutation causes Nance-Horan Syndrome in a Chinese family

Nance-Horan Syndrome (NHS) (OMIM: 302350) was first described in detail by two independent studies in 1974 as a rare X-linked developmental disorder characterized by congenital cataracts, with occasional dental anomalies, brachymetacarpia and mental retardation [1‐4]. It is inherited as an X-linked recessive trait, whereby heterozygous females manifest a similar but milder disease phenotype than affected males [5].

The NHS (Nance-Horan syndrome) gene has been mapped to chromosome Xp22.13 [6]. The gene is composed of 10 coding exons in an approximately 650-kb [7] region, encoding at least four isoforms as a result of alternative splicing. The two major isoforms, NHS-A and NHS-1A, are transcribed from exon 1 and comprise 1630 amino acids and 1651 amino acids, respectively. NHS-B, a 1335-amino acid protein, is transcribed from exon 1b; NHS-C, a 1453-amino acid protein, is transcribed from exon 1a [8‐10].

To date, at least 33 mutations [11‐15], including 6 from the Chinese population, have been reported in the NHS gene, many of which are nonsense mutations or InDels, with few copy number variations and missense and splice site mutations. In this study, we report the identification of a novel splice site mutation in the NHS gene in a Chinese family with NHS. This report broadens the spectrum of NHS gene mutations implicated in NHS pathogenesis.

The study presented here describes the clinical and molecular genetic analysis of a Chinese family exhibiting Nance-Horan Syndrome (NHS). Mutations in the NHS gene have been reported from various populations throughout the world, including Turkish, Indian, Tunisian, Australian, Taiwanese, European and Chinese populations. Clinical evaluation of the available affected male subjects revealed typical NHS ocular and nonocular features that are more severe than in obligate carriers, in concordance with previous reports [8, 11, 17]. Both affected male subjects experienced profound vision loss, and one (II: 5) underwent cataract surgery. Mild mental retardation was exhibited by an affected male (III: 1) in this family. Mental retardation in NHS is highly variable and has been reported in 20–30% of affected subjects, which suggests it is a major component of the NHS syndrome [5].

NHS carrier females manifest mild and variable phenotypes. In one study, Y-sutural posterior lens opacities associated with microcornea and dental anomalies was observed in 100% of carrier females [18, 19], though carriers presenting clear to total lens opacities with small cornea, strabismus and nystagmus have also reported [11]. In our carrier females (II: 2; III: 7), cataract was no longer observed due to cataract surgery prior to the study. Although both carriers were of normal intellect, with normal facial features, microcornea, nystagmus, high myopia and strabismus were found.

Splice site mutations represent a significant proportion of gene alterations leading to Mendelian disorders [20]. Such mutations influencing precursor mRNA splicing either result in complete skipping of one or more exons, retention of introns, creation of a pseudo-exon or activation of a cryptic splice site with in an exon or an intron [21]. Activation of cryptic splice site is the second most frequent consequence of splicing defect after exon skipping [22]. The identified sequence variant (c.1045 + 2T > A) in the NHS gene, located just after exon 4 at the 5′ consensus donor site, disrupts the natural splicing of exons 4-5.

The female fetus was at the 22^nd week of gestation at the time of sample collection for DNA extraction. Sequencing of the NHS gene revealed the splice site mutation in a heterozygous state (m/+). Sonographic scan of the fetus (data not shown) and follow-up ophthalmological examination after birth confirmed the presence of congenital cataract, in concordance with the molecular genetic analyses.

The NHS protein has multiple isoforms, and cell type-dependent differential expression is observed. The two major isoforms, NHS-A and NHS -1A, which are implicated in the pathogenesis of NHS, contain a functional WAVE homology domain at the N-terminus. This domain interacts with members of the Abelson-interactor (Abi) family and contributes to actin remodeling regulation and maintenance of cell morphology [9]. NHS-A, the neuronal and epithelial-specific isoform, is mainly expressed in the human eye lens, and the encoded protein associates with the peripheral cell membrane in the lens epithelium [23]. The identified splice site substitution in intron 4 induces a putative premature stop codon 7 codons downstream (p.G349-IVLVFSX), which is likely to result in a truncated NHS protein. Truncating mutations may lead to nonsense mediated decay (NMD), which results in complete loss of mutated proteins. However, it has been reported that some mRNAs harboring premature termination codons located in some specific regions can escape the NMD pathway, leading to an abnormal protein product [24]. Previously described RT-PCR analysis for a truncating mutation in exon 1(c.115C > T; Q39X) of the NHS gene in an Indian NHS family found no evidence of NMD pathway involvement [17]. In the present study, RT-PCR analysis depicting the presence of an aberrant mRNA transcript in the affected and carrier subjects also suggested escape from the NMD pathway. As a possible consequence, a truncated NHS protein composed of 355 amino acids with a damaging effect would be produced in the cells of these individuals, causing the disease phenotype. However, additional work is required to understand the exact pathogenic mechanism underlying NHS.