GSE126171
The pathognomonic FOXL2 C134W mutation alters DNA binding specificity
The somatic missense point mutation c.402C>G (p.C134W) in the FOXL2 transcription factor is pathognomonic for adult-type granulosa cell tumours (AGCT) and a diagnostic marker for this tumour type. However, the molecular consequences of this mutation and its contribution to the mechanisms of AGCT pathogenesis remain unclear. To explore the mechanisms driving FOXL2C134W pathogenicity we engineered V5-FOXL2 and V5-FOXL2C134W doxycycline inducible isogenic cell lines and performed ChIP-seq and transcriptome profiling following transgenic induction. Interestingly we found that FOXL2C134W associates with the majority of FOXL2 binding sites as well as a large collection of novel regions throughout the genome. De novo motif analysis of the DNA elements uniquely bound by FOXL2C134W revealed a novel DNA motif that was not seen in the shared or FOXL2 specific DNA elements and we validated an altered DNA binding specificity for FOXL2C134W using in vitro band shift assays. Integrated analysis of FOXL2C134W specific proximal promoter binding sites and differential gene expression identified direct targets of FOXL2C134W. We validated one such target and demonstrated a FOXL2C134W dependent sensitivity to a therapeutic that targets survivin (YM155). Taken together our results suggest that FOXL2C134W pathogenicity in AGCT is driven in part by an alteration in DNA binding specificity contributing to an oncogenic transcriptional program. Overall design: Examination of wildtype and mutant FOXL2 DNA binding, gene expression and epigenetic profiling (H3K4me1, H3K4me3, H3K27me3, H3K27ac) in SVOG3e inducible cell lines using massively parallel sequencing-based assays.