The epithelial splicing regulatorESRP2is epigenetically repressed by DNA hypermethylation in Wilms tumour and acts as a tumour suppressor

Abstract

Wilms tumour (WT), a childhood kidney cancer with embryonal origins, has been extensively characterised for genetic and epigenetic alterations, but a proportion of WTs still lack identifiable abnormalities. To uncover DNA methylation changes critical for WT pathogenesis, we compared the epigenome of fetal kidney with two WT cell lines, using methyl-CpG immunoprecipitation. We filtered our results to remove common cancer-associated epigenetic changes, and to enrich for genes involved in early kidney development. This identified four candidate genes that were hypermethylated in WT cell lines compared to fetal kidney, of which ESRP2 (epithelial splicing regulatory protein 2), was the most promising gene for further study. ESRP2 was commonly repressed by DNA methylation early in WT development (in nephrogenic rests) and could be reactivated by DNA methyltransferase inhibition in WT cell lines. When ESRP2 was expressed in WT cell lines, it acted as an inhibitor of cellular proliferation in vitro and in vivo it suppressed tumour growth of orthotopic xenografts in nude mice. RNA-seq of the ESRP2-expressing WT cell lines identified several novel splicing targets, in addition to well-characterised targets of ESRP2. One of these targets, LEF1, is a component of the Wnt signalling pathway that is essential for kidney development and commonly disrupted in WT. We propose a model in which the Wnt pathway can be disrupted in early kidney development to generate WT, either by genetic abnormalities such as WT1 mutations, or by epigenetic defects, such as ESRP2 methylation.

Transcriptomic Analyses of MYCN-Regulated Genes in Anaplastic Wilms’ Tumour Cell Lines Reveals Oncogenic Pathways and Potential Therapeutic Vulnerabilities

“…The Wit49 [18] and 17.94 [19] anaplastic WT cell-lines were kind gifts from Prof. Herman Yeger and Dr. Keith Brown, respectively. The identity of both cell-lines was confirmed by short tandem repeat (STR) analysis [19,20]. Wit49 cells were cultured at 37 • C under 5% CO 2 in Dulbecco's modified Eagle's medium supplemented with 15% fetal calf serum, 2 mmol/L L-glutamine, 0.1 mg/mL penicillin/streptomycin, 0.6% (v/v) β-mercaptoethanol and 1× insulin-transferrin-selenium, all purchased from Sigma.…”