Abstract
Purpose :
Blepharophimosis, ptosis, and epicanthus inversus syndrome (BPES) is a rare autosomal dominant condition involving malformations of the eyelid with additional features. FOXL2 gene mutations account for 72% of BPES cases. We report a multi-generational Ashkenazi Jewish (AJ) family with 4 individuals with classical BPES, lateral eyebrow dystopia, eyelid hirsutism, ciliated and keratinized conjunctiva, and absent eyelashes. Sanger sequencing and locus marker analysis excluded a causative FOXL2 mutation. Exome sequencing was used to identify the gene defect causing this novel BPES-like phenotype.
Methods :
Exome sequencing using the Nimblegen SeqCap EZ V3 capture kit and 2x100bp reads on an Illumina HiSeq2000 was undertaken on 6 family members (4 affected and 2 unaffected). Variants were filtered and analyzed using Golden Helix SVS software. The ExAC database, 162 unrelated exomes, and ethnically matched controls from the Ashkenazi Genome Consortium were used to filter out common variants and calculate rare variant allele frequencies. In silico tools were used to assess the degree of evolutionary conservation at variant residues (GERP++, phyloP), and predict pathogenicity (SIFT, Polyphen2, Mutation Taster). Sanger sequencing validated candidate variants and enabled co-segregation analysis. Immunohistochemistry (IHC) was performed on P14 mouse eyelid tissue using a Peptidase D (PEPD) antibody to assess protein expression.
Results :
Exome sequencing identified a short list of 45 co-segregating rare variants, from which a novel heterozygous c.853G<A (p.G285S) missense variant was seen in PEPD. The variant allele was not present in 128 AJ and >20,000 individuals globally. The glycine residue has been conserved for 450 million years, since pufferfish and humans shared a common ancestor. In silico tools predicted the variant to be damaging. IHC showed PEPD expression in eyelid, and especially hair follicles.
Conclusions :
We have identified a novel variant in PEPD which may cause a BPES-like phenotype. The low allele frequency, strong functional conservation, and expression pattern in eyelid tissue provide compelling evidence. Furthermore, loss-of-function mutations in PEPD have been associated with facial dysmorphism, ptosis, hypertelorism, and hirsutism in patients with recessively inherited prolidase deficiency. Additional functional characterization of this variant is underway using cellular and animal model systems.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.