Conservation of cis-regulatory codes over half a billion years of evolution

The identification of homologous cell types across species represents a crucial step in understanding cell type evolution. The retina is particularly amenable to comparative analysis because the basic morphology, connectivity, and function of its six major cell classes have remained largely invariant since the earliest stages of vertebrate evolution. We used comparative single-nucleus chromatin accessibility analysis of lamprey, fish, bird, and mammalian retinas, which began to diverge over half a billion years ago, to demonstrate cross-species conservation of cis-regulatory codes in all six retinal cell classes. In this study, we acquired retinal single-cell gene expression profiling (scRNA-seq) and single-nucleus chromatin accessibility (snATAC-seq) from sea lamprey (Petromyzon marinus), a jawless species. We also acquired long-read sequence data from lamprey retina and single-nucleus chromatin accessibility from chicken (Gallus gallus). A publicly available sea lamprey transcriptome assembly was updated using newly generated long-read sequencing data. Subsequently, single nuclei multiome profiles (comprising RNA and ATAC) of the downstream migrant lamprey retina were generated with the updated assembly. Furthermore, a single-nuclei chromatin accessibility profile was generated from the newly hatched (P0) chicken, which serves as a comparison of cis-regulatory features across diverse vertebrate species.