Potential involvement of abnormal splicing in severe WT1-related disorders

WT1-related disorders are associated with WT1 gene variants. Recent advances in genetic medicine have led to a better understanding of the genotype-phenotype correlation in WT1-related diseases, particularly missense variants in exons 8 or 9 that lead to a wide range of severities. Exonic variants can lead to splicing abnormalities in rare diseases. No reports have investigated exonic variants in WT1 that cause aberrant splicing. We examined whether exonic variants in WT1 exon 8 or 9 cause splicing abnormalities and affect disease severity.

We selected nine rare missense variants in exon 8 or 9 in WT1 outside the DNA binding domain and C2H2 sites from the Human Gene Variant Database Professional that unexpectedly present severe phenotypes. We conducted functional splicing assays using hybrid minigenes for the nine variants containing exon 8 and 9 and surrounding sequences. Minigene vectors were transfected into cultured cells, and mRNA was analyzed. In silico analysis was performed.

Splicing assays revealed that one of the nine variants caused aberrant splicing, with exon 8 skipping. One previously reported case with this variant showed particularly severe phenotype, progressing to kidney failure within 3 months.

One WT1 variant in exon 8 or 9 disrupted the splice site, leading to aberrant splicing in vitro and potentially contributing to an unexpectedly severe phenotype for a missense variant outside the DNA binding domain and C2H2 sites. In vitro splicing assays may help clarify the genotype-phenotype correlation in WT1-related disorders, especially for variants outside canonical functional domains.