Alternative splicing (AS) plays critical roles in controlling normal developmental programs. ESRP1 (Epithelial Splicing Regulatory Protein) is important cell-type specific regulator that affects splicing of various genes. To date, a considerable number of cancer-related genes have been shown to undergo splicing alterations during ovarian and testicular cancer progression. In this study, we have compared ESRP1 expression in human ovarian and testicular, normal and tumor tissues. In addition, potential splicing targets of ESRP1 have been examined in a seminoma-derived cell line (TCam-2).
We examined expression of ESRP1 and its putative targets, (FGFR2 and CD44), in human ovarian and testicular normal and tumor tissues using droplet digital PCR. In addition, RNA interference was used to knockdown Esrp1 expression in TCam-2 followed by transcriptional and splicing analysis using RNA-Seq.
Our results showed upregulation of Esrp1 in both ovarian and testicular cancer and a correlation with switching from mesenchymal to epithelial isoforms of CD44 and FGFR2 in ovarian but not testicular cancers. RNA sequencing of Esrp1 depleted cells resulted in the identification of 576 novel potential splicing targets for ESRP1 in this germ cell tumour cell line. Also, splicing changes in some of these targets genes have been confirmed using RT-PCR. IPA analyses demonstrated ESRP1 regulates alternative splicing of genes that are involved in directing critical pathways involved in cell migration and morphology that occur during EMT. Our data also showed that four mitochondrial complexes of oxidative phosphorylation are affected by differential gene expression after silencing of ESRP1.
Overall, these data suggest that ESRP1 is involved in gonadal cancer progression by regulation of AS of numerous genes that are related to EMT. Furthermore, differential gene expression after silencing of ESRP1 suggests that ESRP1 expression in testicular germ cells may alter ATP production and affect energy metabolism of these cells.