Comprehensive single cell transcriptome lineages of a proto-vertebrate

Studies of ascidian (sea squirt) embryos have highlighted the importance of cell lineages in animal development for over 100 years. As simple proto-vertebrates, they are also used to explore the evolutionary origins of novel cell types, such as cranial placodes and neural crest in vertebrates. To build upon these efforts we have determined comprehensive single cell transcriptomes of Ciona intestinalis throughout embryogenesis. More than 90,000 cells from 10 different developmental stages were examined, spanning the entirety of morphogenesis, from the onset of gastrulation at the 110-cell stage to the hatching of swimming tadpoles. This represents an average of over 12-fold coverage for every cell at every stage of development, owing to the small cell numbers that are a hallmark property of ascidian embryogenesis. Single cell transcriptome trajectories were used to construct “virtual” cell lineage maps, which confirm and extend those determined by conventional labeling methods. These datasets were also used to reconstruct regulatory cascades and provisional gene networks for a variety of cell types, including nearly 40 different neuronal subtypes comprising the larval nervous system. We summarize several applications of these datasets, including the identification of individual transcriptomes within the complete synaptome of swimming tadpoles, visualizing dynamic changes in gene expression during the birth, migration, and axogenesis of defined neurons, and the evolution of novel cell types such as the vertebrate telencephalon. Overall design: Synchronized embryos from 10 different stages of development were rapidly dissociated in RNase-free calcium-free synthetic seawater, and individual cells were processed in the 10x Genomics Chromium system with at least 2 biological replicates for each developmental stage. The staged embryos span all of the hallmark processes of development, beginning with gastrulation to swimming tadpoles when all larval cell types, tissues and organs are formed. In total, we profiled 90,579 cells, corresponding to an average of over 12-fold coverage for every cell across each of the sampled stages

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Cao C, Lemaire LA, Wang W, Yoon PH, Choi YA, Parsons LR, Matese JC, Wang W, Levine M, Chen K