Comparative single-cell analyses reveal evolutionary repurposing of a conserved gene program in bat wing development [ChIPseq]

Bats (order Chiroptera) are the only mammals capable of self-powered flight. Unlike in humans and mice, digits II-V in the bat forelimb are not separated but connected by the chiropatagium, an elastic membrane that forms the wing. The molecular underpinnings of these morphological differences, one of the most striking adaptations in mammalian evolution, remain obscure. Here, we use a suite of omics tools and single-cell analyses to compare the development of fore- and hindlimbs, in bat and mouse. We demonstrate a clear conservation of cell states and processes between species, even at the level of apoptosis, a mechanism that removes the tissue between the digits. To trace down the cellular origin of the persistent interdigital cells in bats, we micro-dissected the embryonic chiropatagium and performed single-cell transcriptomics. We found that the chiropatagium cells develop from fibroblasts populations, and independently from the apoptosis-related retinoic acid signaling cells of the interdigital mesenchyme. These cells from the distal part of the limb repurpose a gene program otherwise only present in proximal cells. Epigenomic and gene network analyses revealed two transcription factors, MEIS2 and TBX3, among the most prominent regulators of this gene program. Transgenic ectopic expression of MEIS2 and TBX3 in interdigital cells of mice resulted in the activation of genes associated with bat wing processes. Major morphological changes in these mutants include an increase in autopod volume, extracellular matrix and the partial retention of interdigital tissue with fusion (syndactyly) of digits. Our results elucidate fundamental molecular mechanisms of wing formation in bats. More generally, we illustrate that the repurposing of existing developmental programs is an evolutionary molecular mechanism to generate morphological novelties. Overall design: We used ChIP-seq against H3K27ac histone modification to identify active regulatory elements in developing limbs. We used ChIP-seq against Meis2 to detect bound downstream targets of this transcription factor. *************************************************************** The table below lists GEO accessions reused/reanalyzed for this study. Processed data file(s): Meis2_mm39_forelimb_E105.narrowPeak ***************************************************************