SRP408920 PRJNA903839 GSE218437 Co-option of the transcription factor SALL1 in mole ovotestis formation [4C] Changes in gene expression represent an important source for phenotypical innovation. Yet, how such changes emerge and impact the evolution of traits remains elusive. Here, we explore the molecular mechanisms associated with the development of masculinizing ovotestes in female moles. By performing comparative analyses of epigenetic and transcriptional data in mole and mouse, we identified SALL1 as a co-opted gene for the formation of testicular tissue in mole ovotestes. Chromosome conformation capture analyses highlight a striking conservation of the 3D organization at the SALL1 locus, but a prominent evolutionary turnover of enhancer elements. Interspecies reporter assays support the capability of mole-specific enhancers to activate transcription in urogenital tissues. Through overexpression experiments in transgenic mice, we further demonstrate the capability of SALL1 to induce the ectopic gene expression programs that are a signature of mole ovotestes. Our results highlight the co-option of gene expression, through changes in enhancer activity, as a prominent mechanism for the evolution of traits. Overall design: We used 4C-seq with the viewpoint in the Sall1 promoter to measure the interacctions of the gene with its regulatory landscape GSE218437 pubmed 37519269 parent_bioproject PRJNA906599