Fig. 5

FGF-MEK1/2 signalling supports expression of stem cell-associated genes in early germ cells, but only MEK1/2 signalling is required for male germline differentiation. RNA sequencing analysis of germ cells from XX or XY E12.5 gonads, or XX or XY E12.5 gonads cultured for 24 or 72 h with DMSO or 500 nM FGFRi or MEKi. A Example of FACS scatterplot depicting GFP-positive germ cell isolation. B Number of differentially expressed genes (DEGs) between XX or XY E12.5 (time 0) and XX or XY DMSO controls from 24 and 72 h cultures. C Multidimensional scaling (MDS) of all control conditions. D MDS of XX and XY gonads cultured for 24 h. E Differential gene expression analysis of XY E12.5 + 24 h FGFRi XY (i) or XY E12.5 + 24 h MEKi (ii) vs XY E12.5 + 24 h DMSO. F Venn diagram of 24 h FGFRi and MEKi DEGs (i) and heatmap of common DEGs (ii). Asterisks represent genes associated with germ cell tumours, cancer/stem cell biology and/or proliferation. G MDS of XX and XY gonads cultured for 72 h. H Differential gene expression analysis of XY E12.5 + 72 h FGFRi (i) or XY E12.5 + 72 h MEKi (ii) vs XY E12.5 + 72 h DMSO. I Venn diagram comparing MEKi 72 h culture DEGs expressed lower (i) or higher (ii) than expected with XX or XY specific genes identified in B. J Heatmap of DEGs identified in XY E12.5 + 72 h MEKi vs XY E12.5 + 72 h DMSO associated with male germline differentiation, DNA methylation/piRNA pathway, cell cycle, female germline differentiation/meiosis, Nodal signalling, pluripotency and spermatogonial stem cells (SSCs). Asterisks highlight genes associated with cell cycle, meiosis and pluripotency but are not differentially expressed. For all comparisons, genes with FDR < 0.05 and |logFC|> 0.585 (equivalent to |FC|> 1.5) were considered differentially expressed