A novel frameshift mutation in FRMD7 causes X-linked infantile nystagmus in a Chinese family

A Chinese family with idiopathic infantile nystagmus (IIN) was recruited at the Department of Ophthalmology in XinHua hospital, Shanghai Jiao Tong University. All patients received comprehensive ophthalmic examinations, and eye movements were recorded. Then,5-ml of blood samples were collected to conduct high-throughput sequencing. This study was approved by the Ethics Committee of the XinHua hospital and conformed to the tenets of the Declaration of Helsinki. Informed consent was obtained from all participants. (If the patient was a minor, the informed consent was signed by parents or guardian)

A comprehensive ophthalmic examinations was performed on each subject. We used Snellen visual acuity charts and the Titmus test to measure visual acuity and stereopsis. Visual acuity was measured monocularly and binocularly for distance both at the patient’s AHP and corrected posture with the patient’s best optical correction in place. The Hirschberg test and the prism cover test were used to assess for strabismus. We used a divider with a linear calibration (goniometer) to measure the degree of AHP when patients looked at a target from 5-m away. One arm of the goniometer was in the sagittal direction of the head. The other arm was in the sagittal direction of the body. The angle of the corner of the two arms defined the angle of the AHP. Horizontal and vertical eye-movement measurements were taken using IR reflection with a sampling frequency of 500 Hz. Calibration was performed with the opposite eye covered. The patient was required to fixate horizontally from 0 to 30° in 5-degree increments and vertically from 0 to 20° both monocularly and binocularly. The position of the null zone was determined by eye movement recordings. The duration of each examination was approximately 10 seconds. Ocular examinations also included cycloplegic refraction, slit-lamp examination, electroretinography and visual evoked potential response. From fundus photographs, we analyzed each patient’s retinal vasculature, retinal pigment and the morphology of the optic nerve head, including the peripapillary double ring sign. We also measured the cup-to-disc ratio (C/D ratio) and the disc–macula distance to disc diameter ratio (DM:DD ratio). When the DM:DD ratio was greater than 3.0, a small optic disc could be indicated.

Mutations in the FRMD7 gene were screened by high-throughput sequencing using genomic DNA from the patient’s peripheral blood. We used a DNA extraction kit (TIANGEN, Beijing, China) according to the manufacturer’s instructions. DNA was quantified using a Nanodrop 2000 PCR-amplification of all of FRMD7’s all coding exons and intron-exon junctions was performed with the primers according to a previous report [5]. PCR products were sequenced on an ABI PRISM 3730XL DNA analyzer (Applied Biosystems, Foster,CA,USA). The sequences were aligned, and the mutations were detected using mutation surveyor software (Softgenetic,Pennsylvania,USA). The effects of the mutations on the protein coding region (synonymous, missense, nonsense, frameshift, etc.) were predicted by Exome-assistant, and the potential pathogenic effects of the mutations on protein function were estimated using Mutation Taster (http://​www.​mutationtaster.​org), Polyphen-2 (http://​genetics.​bwh.​harvard.​edu/​pph2/​) and SIFT (http://​sift.​jcvi.​org/​www/​SIFT_​enst_​submit.​html). The database we used were the ExAC (http://​exac.​broadinstitute.​org) ,the exome Variant Server (http:// evs.gs.washington.edu/EVS/), and the 1000 Genomes Project (http://​www.​1000genomes.​org/​).

There were 20 people in this Chinese family; 10 of them had IN (5 female and 5 male). All IN patients we examined presented nystagmus within the first six months of life, conjugate horizontal oscillation of the eyes, reduced visual acuity and stereopsis. The mean LogMAR binocular visual acuity measured at both their AHP and corrected head posture was 0.25 and 0.37, respectively. No patient had associated strabismus. Four patients had obvious AHPs and two chose to receive surgery to shift the null zone. Four had similar nystagmus wave forms(bidirectional jerk, BDJ). One patient had pure jerk nystagmus (J), jerk nystagmus with extended foveation (JEF) and bidirectional jerk nystagmus (BDJ). Another had pseudo-pendular nystagmus with foveating saccades (PPFS). None had anterior-segment pathologies or macular hypoplasia. The mean C/D ratio was 0.35± 0.14 (right eye) and 0.39± 0.10 (left eye). The mean DM/DD ration was 2.66± 0.77 (right eye) and 2.43± 0.68 (left eye). No one had obvious morphological changes of the optic nerve head. However, one patient’ s DM:DD ratio was greater than 3.0.

Sequence analysis of this Chinese family detected a novel frameshift mutation c.823-829delACCCTAC in the 9th exon of FRMD7. This mutation results in a frameshift mutation at codon 275 (p.Thr275fs) and predicts protein truncation. This mutation was confirmed to extend to other affected family members. However, this mutation was not found in the unaffected members or in the 100 unrelated controls (Fig. 1). There was no male-to-male transmission, female carriers were heterozygous, and male carriers were hemizygous. All male carriers were affected. The genetic pattern of disease conforms to X-linked dominant inheritance (Table 1 and Fig. 2). Some female members (III-2, III-5) heterozygous for the FRMD7 mutation did not have IN. This finding suggested incomplete penetrance for this truncating FRMD7 mutation.