Evolution of the regulation of developmental gene expression in blind Mexican cavefish

Changes in gene expression regulation during development are considered the main drivers of morphological evolution and diversification. Here, we analysed the embryonic transcriptomes of surface-dwelling and blind cave-adapted morphs of the species Astyanax mexicanus and their reciprocal F1 hybrids at tailbud stage. Comparing gene expression in parents and allelic expression ratios in hybrids, we found that [~]20% of the transcriptome is differentially expressed and that cis-regulatory changes are the main contributors to variations in early developmental gene expression in the two morphs. We provide a list of 108 cis-regulated genes that could contribute to cavefish developmental evolution, and further explore the regulatory mechanisms controlling the cellular and regional expression of rx3, a "master eye gene". Using quantitative embryology approaches after fluorescent in situ hybridisation, cell transplantations and interference with signalling pathways, we show that rx3 cellular levels -controlling optic cell fates-are regulated in cis and in a cell-autonomous manner, whereas the size of rx3 domain -controlling eye size-depends on non-autonomous Wnt signalling. Altogether, we reveal how distinct mechanisms and regulatory modules can regulate developmental gene expression and shape developmental evolution, with negligible contribution of coding mutations.