A 127 kb truncating deletion of PGRMC1 is a novel cause of X-linked isolated paediatric cataract

Abstract

Inherited paediatric cataract is a rare Mendelian disease that results in visual impairment or blindness due to a clouding of the eye's crystalline lens. Here we report an Australian family with isolated paediatric cataract, which we had previously mapped to Xq24. Linkage at Xq24-25 (LOD = 2.53) was confirmed, and the region refined with a denser marker map. In addition, two autosomal regions with suggestive evidence of linkage were observed. A segregating 127 kb deletion (chrX:g.118373226_118500408del) in the Xq24-25 linkage region was identified from whole-genome sequencing data. This deletion completely removed a commonly deleted long non-coding RNA gene LOC101928336 and truncated the protein coding progesterone receptor membrane component 1 (PGRMC1) gene following exon 1. A literature search revealed a report of two unrelated males with non-syndromic intellectual disability, as well as congenital cataract, who had contiguous gene deletions that accounted for their intellectual disability but also disrupted the PGRMC1 gene. A morpholino-induced pgrmc1 knockdown in a zebrafish model produced significant cataract formation, supporting a role for PGRMC1 in lens development and cataract formation. We hypothesise that the loss of PGRMC1 causes cataract through disrupted PGRMC1-CYP51A1 protein-protein interactions and altered cholesterol biosynthesis. The cause of paediatric cataract in this family is the truncating deletion of PGRMC1, which we report as a novel cataract gene.

Fig. 1. Pedigree of family CRVEEH66 and candidate variant status.

The previously studied CRVEEH66 family [12]. An ophthalmologist confirmed cataract affection status and no clinically significant lens opacity was observed in females. Segregation of 127 kb deletion (Del), at Xq24, is displayed. All affected males are hemizygous for the deletion and obligate female carriers are heterozygous for the variant (Del/WT) as is female IV:14.

Fig. 2. CRVEEH66 genome-wide parametric linkage analysis results.

Multipoint LOD scores (y-axis), indicating evidence for linkage, are displayed across the genome (x-axis). A linkage peak on chromosome X (Chr. 23), between markers rs2428312 and rs7887767, reaches significance with a maximum LOD score of 2.53. Regions on both chromosome 1 (rs696859–rs1316440) and chromosome 3 (rs4527385–rs4857688) reached maximum LOD scores of 2.44, with autosomal LOD scores exceeding two suggestive evidence of linkage (dashed line). The cut-off value for significance is indicated by the red line.

Fig. 3. CRVEEH66 127 kb Xq24 deletion variant validation.

A Agarose gel electrophoresis results of representative samples for PCR products: deletion, left breakpoint, and right breakpoint. Heterozygous female obligate carrier and hemizygous affected male both possess the deletion amplicon. Unaffected female and male samples display wild-type results, with amplicons for breakpoints only. M; 100 bp ladder, C; PCR no template control. B Diagrammatic representation of the 127 kb deletion, indicated by the hatched region. Sequencing chromatograms at the critical breakpoints show consensus with the related halves of the deletion sequence. Deletion PCR fragment displayed from sequencing with the reverse primer. C UCSC genome browser displays the whole-genome sequence coverage from III:12 in dark red compared to an unrelated control sample in blue focused on the region surrounding the PGRMC1 gene. The coverage peaks within the deletion region (as indicated by the marked drop in coverage) in individual III:12 correspond with repeat regions in the genome (data not shown) and likely represent spurious mapping.

Fig. 4. Zebrafish pgrmc1 morpholino-induced knockdown lens phenotypes.

Representative DIC lens images from pgrmc1 morpholino experiments. Wild-type Gal4^s1020t/UAS:mCherry transgenic zebrafish display healthy lenses (A). Control fish injected with standard control morpholino display healthy lens (B) and minor cataract with very fine nuclear pitting (C). Fish injected with pgrmc1_MO1 (2 ng) morpholino displayed both healthy lenses (D) or cataracts of variable severity; minor nuclear central mass (E), mild nuclear central mass with fibre cell disorganisation (F) and moderate/severe nuclear density with pitting (G). Fish injected with pgrmc1_MO2 (12 ng) morpholino displayed mainly healthy lenses (H) or subtle cataracts of variable severity; minor distinct central lens density (I), minor nuclear fibre cell disorganisation (J) and moderate nuclear fibre cell disorganisation with minor pitting (K).