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Unexpected ABCC6 mRNA splicing in a Chinese family with pseudoxanthoma elasticum
Author(s) -
Sun Qiao,
Wang Weijun,
Wu Lifang,
Cheng Lu,
Tong Xuemei,
Xu Xun
Publication year - 2019
Publication title -
acta ophthalmologica
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.534
H-Index - 87
eISSN - 1755-3768
pISSN - 1755-375X
DOI - 10.1111/aos.13819
Subject(s) - minigene , rna splicing , exon skipping , splice site mutation , exon , pseudoxanthoma elasticum , intron , genetics , biology , sanger sequencing , alternative splicing , mutation , microbiology and biotechnology , medicine , gene , rna , pathology
Purpose To identify the clinical characteristics and pathogenic genes among a Chinese family with angioid streaks and to assess a novel splicing mutation at the transcriptional and translational levels. Methods Consenting family members were clinically evaluated, and blood samples were collected for targeted exome capture sequencing and/or Sanger sequencing. The two affected siblings were assessed by multimodal fundus imaging. ABCC6 splicing patterns were analysed by RNA identification and quantification using the proband's peripheral blood mononuclear cells. Minigene experiments were performed to verify the university. Plasmids expressing the transcripts were transfected into HEK293 cells to assess protein translation. Bioinformatic analyses were also performed to predict the splicing patterns and the functional consequences of the mutation. Results The two siblings were trans‐compound heterozygous pseudoxanthoma elasticum (PXE) patients with the same genotype (c.3703C>T and c.1177‐2A>G for ABCC6 ) but different phenotypes. We identified several ABCC6 alternative splicing transcripts that were not previously reported. The novel splicing mutation c.1177‐2A>G led to the upregulation of three transcripts, one using a cryptic splice acceptor in the upstream region of the intron, another using a cryptic splice acceptor in the downstream exon, and the third stimulating non‐canonical U12‐type splicing. All the transcripts were successfully translated in vitro . Conclusion The genotype–phenotype correlation of PXE is poorly understood. The novel ABCC6 splicing mutation c.1177‐2A>G results in multiple splicing patterns. Endogenous U2 to U12 conversion may occur in humans in a disease state. Peripheral blood mononuclear cells can be reliably used to study ABCC6 RNA.