The ability to profile hundreds of thousands to millions of single cells using scRNA-sequencing has revolutionized the fields of cell and developmental biology, providing incredible insights into the diversity of forms and functions of cell types across many species. These technologies hold the promise of developing detailed cell type phylogenies which can describe the evolutionary and developmental relationships between cell types across species. This will require sampling of many species and taxa using single-cell transcriptomics, and methods to classify cell type homologies and diversifications. Many tools currently exist for analyzing single cell data and identifying cell types. However, cross-species comparisons are complicated by many biological and technical factors. These factors include batch effects common to deep-sequencing approaches, well known evolutionary relationships between orthologous and paralogous genes, and less well-understood evolutionary forces shaping transcriptome variation between species. In this review, I discuss recent developments in computational methods for the comparison of single-cell-omic data across species. These approaches have the potential to provide invaluable insight into how evolutionary forces act at the level of the cell and will further our understanding of the evolutionary origins of animal and cellular diversity.

Highlights d Chromosomes from early embryos resemble a barbell d Lamina interactions stretch chromosomes and separate compartments d Conventional compartments arise during gastrulation via long-distance associations d Single-chromosome clustering uncovers prevalent conformations

Hundreds of cell types form the vertebrate brain, but it is largely unknown how similar these cellular repertoires are between or within species, or how cell type diversity evolves. To examine cell type diversity across and within species, we performed single-cell RNA sequencing of ~130,000 hypothalamic cells from zebrafish ( Danio rerio ) and surface-and cave-morphs of Mexican tetra ( Astyanax mexicanus ). We found that over 75% of cell types were shared between zebrafish and Mexican tetra, which last shared a common ancestor over 150 million years ago. Orthologous cell types displayed differential paralogue expression that was generated by sub-functionalization after genome duplication. Expression of terminal effector genes, such as neuropeptides, was more conserved than the expression of their associated transcriptional regulators. Species-specific cell types were enriched for the expression of species-specific genes, and characterized by the neo-functionalization of members of recently expanded or contracted gene families. Within species comparisons revealed differences in immune repertoires and transcriptional changes in neuropeptidergic cell types associated with genomic differences between surface-and cave-morphs. The single-cell atlases presented here are a powerful resource to explore hypothalamic cell types, and reveal how gene family evolution and the neo-and sub-functionalization of paralogs contribute to cellular diversity.