A splice-site variant in the lncRNA gene RP1-140A9.1 cosegregates in the large Volkmann cataract family

PURPOSE: To identify the mutation for Volkmann cataract (CTRCT8) at 1p36.33. METHODS: The genes in the candidate region 1p36.33 were Sanger and parallel deep sequenced, and informative single nucleotide polymorphisms (SNPs) were identified for linkage analysis. Expression analysis with reverse transcription polymerase chain reaction (RT-PCR) of the candidate gene was performed using RNA from different human tissues. Quantitative transcription polymerase chain reaction (qRT-PCR) analysis of the GNB1 gene was performed in affected and healthy individuals. Bioinformatic analysis of the linkage regions including the candidate gene was performed. RESULTS: Linkage analysis of the 1p36.33 CCV locus applying new marker systems obtained with Sanger and deep sequencing reduced the candidate locus from 2.1 Mb to 0.389 Mb flanked by the markers STS-22AC and rs549772338 and resulted in an logarithm of the odds (LOD) score of Z = 21.67. The identified mutation, rs763295804, affects the donor splice site in the long non-coding RNA gene RP1–140A9.1 (ENSG00000231050). The gene including splice-site junctions is conserved in primates but not in other mammalian genomes, and two alternative transcripts were shown with RT–PCR. One of these transcripts represented a lens cell–specific transcript. Meta-analysis of the Cross-Linking-Immuno-Precipitation sequencing (CLIP-Seq) data suggested the RNA binding protein (RBP) eIF4AIII is an active counterpart for RP1–140A9.1, and several miRNA and transcription factors binding sites were predicted in the proximity of the mutation. ENCODE DNase I hypersensitivity and histone methylation and acetylation data suggest the genomic region may have regulatory functions. CONCLUSIONS: The mutation in RP1–140A9.1 suggests the long non-coding RNA as the candidate cataract gene associated with the autosomal dominant inherited congenital cataract from CCV. The mutation has the potential to destroy exon/intron splicing of both transcripts of RP1–140A9.1. Sanger and massive deep resequencing of the linkage region failed to identify alternative candidates suggesting the mutation in RP1–140A9.1 is causative for the CCV phenotype.