Mutant FOXL2C134W hijacks SMAD4 and SMAD2/3 to drive adult granulosa cell tumors

The FOXL2 mutant C134W occurs in virtually all adult ovarian granulosa cell tumors (AGCT), and is considered a driver of oncogenesis in this disease. However, the mechanism by which FOXL2C134W contributes to tumorigenesis is not known. Here, we show that mutant FOXL2C134W acquires the ability to bind SMAD4, forming a FOXL2C134W/SMAD4/SMAD2/3 complex that binds a novel hybrid DNA motif AGHCAHAA, unique to the FOXL2C134W mutant. This binding induces an enhancer-like chromatin state leading to transcription of nearby genes, many of which are characteristic of stemness and epithelial-to-mesenchymal transition. Importantly, primary AGCT tumors display a strong FOXL2C134W enrichment at hybrid loci. Ablation of SMAD4 or SMAD2/3 inhibits FOXL2C134W binding at hybrid sites and decreases transcription of associated genes. Accordingly, TGFβ-inhibition mitigates the transcriptional effect of FOXL2C134W. FOXL2, SMAD4, SMAD2/3, H3K27ac, K4me1 enrichment profiles (ChIPseq) analyzed in human granulosa cells HGrC1 and KGN in the presence or absence of TGFbeta stimulation. HGrC1 cells express A few samples lack processed data because they were replicates or controls not discussed in the manuscript.