Validation and Classification of Atypical Splicing Variants Associated With Osteogenesis Imperfecta

Abstract

Osteogenesis Imperfecta (OI) is a rare inherited bone dysplasia, which is mainly caused by mutations in genes encoding type I collagen including COL1A1 and COL1A2. It has been well established to identify the classical variants as well as consensus splicing-site-variants in these genes in our previous studies. However, how atypical variants affect splicing in OI patients remains unclear. From a cohort of 867 OI patients, we collected blood samples from 34 probands which contain 29 variants that are located close to splice donor/acceptor sites in either COL1A1 or COL1A2. By conducting minigene assay and sequencing analysis, we found that 17 out of 29 variants led to aberrant splicing effects, while no remarkable aberrant splicing effect was observed in the remaining 12 variants. Among the 17 variants that affect splicing, 14 variants led to single splicing influence: 9 led to exon skipping, 2 resulted in truncated exon, and 3 caused intron retention. There were three complicated cases showing more than one mutant transcript caused by recognition of several different splice sites. This functional study expands our knowledge of atypical splicing variants, and emphasizes the importance of clarifying the splicing effect for variants near exon/intron boundaries in OI.

Case Report: Complete Maternal Uniparental Disomy of Chromosome 2 With a Novel UNC80 Splicing Variant c.5609-4G> A in a Chinese Patient With Infantile Hypotonia With Psychomotor Retardation and Characteristic Facies 2

“…Bioinformatics analysis showed that the variant might be involved in splicing modulation. Similar variants, which lead to protein truncation have been previously reported ( Anna and Monika, 2018 ; Li et al, 2019 ; Śmigiel et al, 2020 ). And in vitro cytology experiments have demonstrated that loss-of-function mutations in UNC80 are associated with IHPRF2 ( Valkanas et al, 2016 ; Wie et al, 2020 ).…”

Novel De Novo TBL1XR1 Variant Causing PIERPONT Syndrome in an Indian Child: A Case Report and Genotype–Phenotype Review of Reported Patients

“…Arginine methylation is a posttranslational modification which can potentially result in altered protein–protein interaction, impacting normal protein function. 12 13 14 15 16 Another possibility is that the variant in concern, being close to the intron–exon boundary, may possibly create aberrant splicing due to the creation of a GT donor splice site. 10 11 12 13 14 It is also food for thought whether similar altered protein–protein interactions involving arginine, in other genes alike, could be an important pathomechanism clue for other cases of syndromic and nonsyndromic autism.…”

Genetic analysis in Japanese patients with osteogenesis imperfecta: Genotype and phenotype spectra in 96 probands

“…Third, we did not perform functional analyses on the proteins, mRNAs, or cDNAs isolated from cultured fibroblasts. Such assays detect quantitative and/or qualitative defects and may help elucidate variants within the clinical spectrum (Li et al, 2019; Van Dijk et al, 2012).…”