The Shh/Gli3 gene regulatory network precedes the origin of paired fins and reveals the deep homology between distal fins and digits

One of the central problems of vertebrate evolution is understanding the relationship among the distal portions of fins and limbs. Lacking comparable morphological markers of these regions in fish and tetrapods, these relationships have remained uncertain for the past century and a half. Here we show that Gli3 functions in controlling the proliferative expansion of distal progenitors are shared among dorsal and paired fins as well as tetrapod limbs. Mutant knockout gli3 fins in medaka (Oryzias latipes) form multiple radials and rays, in a pattern reminiscent of the polydactyly observed in Gli3-null mutant mice. In limbs, Gli3 controls both anterior¿posterior patterning and cell proliferation, two processes that can be genetically uncoupled. In situ hybridization, quantification of proliferation markers, and analysis of regulatory regions reveal that in paired and dorsal fins, gli3 plays a main role in controlling proliferation but not in patterning. Moreover, gli3 down-regulation in shh mutant fins rescues fin loss in a manner similar to how Gli3 deficiency restores digits in the limbs of Shh mutant mouse embryos. We hypothesize that the Gli3/Shh gene pathway preceded the origin of paired appendages and was originally involved in modulating cell proliferation. Accordingly, the distal regions of dorsal fins, paired fins, and limbs retain a deep regulatory and functional homology that predates the origin of paired appendages.This work would have been impossible without the contribution of Jose Luis Gómez-Skarmeta, a beloved friend and mentor whose passion for science was only surpassed by his generosity and kindness. We thank members of the J.L.G.-S. laboratory for fruitful discussions. J.L.G.-S. received funding from the European Research Council under the European Union’s Horizon 2020 research and innovation program (Grant Agreement 740041) and the Spanish Ministerio de Economıa y Competitividad (Grant BFU2016-74961-P) and was also supported, together with J.L.-R. and J.R.M.- M., by institutional grant Unidad de Excelencia Marıa de Maeztu (MDM-2016- 0687, to the Department of Gene Regulation and Morphogenesis of the Centro Andaluz de Biologıa del Desarrollo). J.L.-R. and J.R.M.-M. are supported by the Spanish Ministerio de Ciencia e Innovacion (Grants BFU2017-82974-P PID2020-113497GB-I00 and BFU2017-86339-P/PID2020-112566GB-I00, respectively). N.S. is supported by grants from the Brinson Foundation. J.L. is financed by the Chilean FONDECYT agency (Grant 11180727)