Evolution of cell-type-specific accessible chromatin regions and the cis-regulatory elements that drive lineage-specific innovation

Cis-regulatory elements (CREs) are critical in regulating gene expression, and yet understanding of CRE evolution remains challenging. Here, we constructed a comprehensive single-cell atlas of chromatin accessibility in Oryza sativa, integrating data from 103,911 nuclei representing 126 discrete cell states across nine distinct organs. We used comparative genomics to compare cell-type resolved chromatin accessibility between O. sativa and 57,552 nuclei from four additional grass species (Zea mays, Sorghum bicolor, Panicum miliaceum, and Urochloa fusca). Accessible chromatin regions (ACRs) had different levels of conservation depending on the degree of cell-type specificity. We found a complex relationship between ACRs with conserved noncoding sequences, cell-type specificity, conservation, and tissue-specific switching. Additionally, we found that epidermal ACRs were less conserved compared to other cell types, potentially indicating that more rapid regulatory evolution has occurred in the L1-derived epidermal layer of these species. Finally, we identified and characterized a conserved subset of ACRs that overlapped the repressive histone modification H3K27me3, implicating them as potentially silencer-like CREs maintained by evolution. Collectively, this comparative genomics approach highlights the dynamics of plant cell-type-specific CRE evolution.