Abnormalities in testis development are strongly associated with reproductive health conditions including infertility and testis cancer. In mouse testes, the transcription factor, SOX9 and fibroblast growth factor 9 (FGF9) promote Sertoli cell development, while vascular endothelial growth factor (VEGF) signalling is essential for testicular vasculature. These growth factors signal through downstream pathways such as the MAPK signalling cascade, which is essential for cell proliferation and differentiation, but little is known about MAPK signalling during fetal testis development. We explored MAPK functions in embryonic day (E)12.5 Oct4GFP transgenic mouse testes cultured with MEK1/2 inhibitor for 24 or 72 hours. The MEK1/2 target, phospho-ERK1/2 (pERK1/2), was detected in Sertoli and endothelial cells of controls, but not in MEK1/2 inhibited samples. Flow cytometric and immunofluorescence analyses revealed that MEK1/2 inhibition reduced Sertoli cell proliferation and Sertoli cell localisation to the testis cord basement membrane. RNA sequencing in isolated gonadal somatic cells identified 116 and 114 differentially expressed genes after 24 and 72 hours of MEK1/2 inhibition. Ingenuity Pathway Analysis revealed a striking association of MEK1/2 signalling with angiogenesis, vasculogenesis and cell migration, including a failure to properly express vascular remodelling transcription factors, Sox7 and Sox17, VEGF receptor genes, cell adhesion factor gene, Cd31 and other endothelial markers. Consistent with this, section and wholemount immunofluorescence revealed that CD31, SOX7 and SOX17 protein was lost in MEK1/2 inhibited samples. Similarly, VEGF receptor inhibition reduced CD31, SOX7 and SOX17. Moreover, while VEGF inhibition eliminated pERK1/2 in endothelial cells, pERK1/2 was maintained in Sertoli cells. Our data suggests that VEGF signalling activates MEK1/2 to promote vasculature patterning in the testis and that MEK1/2 signalling independently regulates Sertoli cell proliferation and organisation. As these processes are essential for testis function, this model provides important insights into testicular patterning events that are likely to affect lifelong male reproductive health.