A mutation in SLC24A1 implicated in autosomal-recessive congenital stationary night blindness

Abstract

Congenital stationary night blindness (CSNB) is a nonprogressive retinal disorder that can be associated with impaired night vision. The last decade has witnessed huge progress in ophthalmic genetics, including the identification of three genes implicated in the pathogenicity of autosomal-recessive CSNB. However, not all patients studied could be associated with mutations in these genes and thus other genes certainly underlie this disorder. Here, we report a large multigeneration family with five affected individuals manifesting symptoms of night blindness. A genome-wide scan localized the disease interval to chromosome 15q, and recombination events in affected individuals refined the critical interval to a 10.41 cM (6.53 Mb) region that harbors SLC24A1, a member of the solute carrier protein superfamily. Sequencing of all the coding exons identified a 2 bp deletion in exon 2: c.1613_1614del, which is predicted to result in a frame shift that leads to premature termination of SLC24A1 (p.F538CfsX23) and segregates with the disorder under an autosomal-recessive model. Expression analysis using mouse ocular tissues shows that Slc24a1 is expressed in the retina around postnatal day 7. In situ and immunohistological studies localized both SLC24A1 and Slc24a1 to the inner segment, outer and inner nuclear layers, and ganglion cells of the retina, respectively. Our data expand the genetic basis of CSNB and highlight the indispensible function of SLC24A1 in retinal function and/or maintenance in humans.

Figure 1

Pedigree of Family PKRP070 with Alleles for Chromosome 15q Markers Alleles forming the risk haplotypes are shaded black, alleles cosegregating with the risk haplotype without homozygosity are shaded gray, and alleles not cosegregating with CSNB are shown in white.

Figure 2

Fundus Photographs of Family PKRP070 (A) OD and OS of affected individual 14. (B) OD and OS of affected individual 18. (C) OD and OS of affected individual 28. (D) OD and OS of affected individual 34. (E) OD and OS of unaffected individual 35. OD, oculus dexter (right eye); OS, oculus sinister (left eye).

Figure 3

Sequence Chromatogram Illustrating the Causal Lesions and the Topological Model of SLC24A1 (A–C) Individual 11 (A), individual 12 (B), and individual 14 (C) show the wild-type allele, heterozygous, and homozygous 2 bp deletion c.1613_1614del, respectively. The underlined are the two bases deleted in the affected individuals of family PKFP070. (D) Transmembrane domains of SLC24A1 predicted by Simple Modular Architecture Research Tools (SMART) algorithms. The deletion mutation resides in the fourth transmembrane that constitutes the first of two proposed ion exchanger domains of SLC24A1.

Figure 4

Quantitative Expression Analyses in the Adult Mouse Eye Tissue (A) Expression of Slc24a1 and Rhodopsin was determined by qRT-PCR in the mouse eyes at P01, P03, P05, P10, P15, and P30. Slc24a1 expression is presented as bars on an arbitrary scale, and Rhodopsin expression is presented as a line. All values are the mean (± SEM) of three independent observations after normalization with the control gene (Rpl-19) expression. (B) Quantitative expression of Slc24a1 in adult mouse eye tissue (180 days). Slc24a1 expression is presented as bars using an arbitrary scale on the y axis. Values are presented as the mean (± SEM) of three independent observations after normalization with the control gene (Rpl-19) expression. RPE, retinal pigment epithelium; ON, optic nerve.

Figure 5

Localization of SLC24A1 in Human Retina with In Situ Hybridization Using Digoxigenin-Labeled Sense and Antisense Probes Hybridization with sense probe as a control (left) and antisense probe (right) show strong labeling (black arrows) in the inner segment, outer and inner nuclear layers, and ganglion cell layer. OS, outer segment; IS, inner segment; ONL, outer nuclear layer; OPL, outer plexiform layer; IPL, inner plexiform layer; GCL, ganglion cell layer.

Figure 6

Immunolocalization of Slc24a1 Expression in P13 and Adult Mouse Retinal Layers The slides were stained for Slc24a1 with goat anti-rabbit IgG conjugated with AlexaFluor 594 (red), F-actin using AlexaFluor488 conjugated phalloidin (green), and nuclei using Hoechst 33342 (blue). Merge represents an overlay of images from the first three columns in P13 and adult mouse retinal layers, respectively. The localization pattern illustrates a strong signal in the inner segment, outer and inner nuclear layers, and ganglion cell layer. The scale bar represents 50 um. OS, outer segment; IS, inner segment; ONL, outer nuclear layer; OPL, outer plexiform layer; IPL, inner plexiform layer; GCL, ganglion cell layer.