Spatially-resolved single-cell atlas of ascidian endostyle provides insights into the origin of vertebrate pharyngeal organs
The pharyngeal endoderm, an innovation in deuterostome ancestors, functions as a pharyngeal developmental major player that orchestrate the formation of different germ layers and multiple pharyngeal derivatives in vertebrates. However, the evolutionary origin of multiple pharynx organs in vertebrates remains largely unknown. The endostyle, a distinct pharyngeal organ exclusively presents in basal chordates, represent a good model in understanding the origin of pharyngeal organs. Here, utilizing cutting-edged Stereo-seq and single-cell RNA-seq, we constructed the first spatially-resolved single-cell atlas in the endostyle of ascidian, Styela clava. The spatial location of Stereo-seq and high capture efficiency of single-cell RNA-seq complement each other to ensure the high quality of the atlas. We comprehensively resolved the cellular composition of hemolymphoid region (HLR), which harbors immune and blood cell lineages, illuminating a mixed ancestral structure for blood and lymphoid system. Additionally, we discovered a hair cell-like cell (HCLC) cluster in zone 3, which shares homologous features with vertebrate acoustico-lateralis system hair cells, and the results were further validated by in situ hybridization of cell-specific markers, electronic microscopy and cross-species comparison. These findings reshape the understanding of the pharynx of basal chordate and provide clues for the evolutionary origin of multiplexed pharyngeal organs.