Large-scale identification and analysis of cis-RCNEs in the whole genomes of Brassicaceae

Cis-regulatory conserved noncoding elements ( cis -RCNEs) are non-coding genomic sequences that do not encode rRNA, tRNA, or proteins but exhibit strong evolutionary conservation. These elements are widespread across genomes and play crucial roles in gene expression regulation, influencing species development, phenotypic variation, and adaptive evolution. In this study, we identified 12 498 cis -RCNEs across the whole genomes of 29 Brassicaceae species. Functional enrichment analysis of genes adjacent to these elements revealed a predominant association with growth related and stress-responsive pathways. Using ChIP-seq data, we detected 432 531 regulatory peaks, predominantly localized in promoter regions, particularly within < 1 kb upstream of transcription start sites, with high occupancy rates in Arabidopsis thaliana (> 81%), Brassica rapa (> 47%), Brassica oleracea (> 69%), and Brassica napus (> 64%). Integrative analysis of cis -RCNE target genes and ChIP-seq profiles demonstrated that whole-genome duplication (WGD) derived genes represent a substantial fraction of duplicated genes and exhibit heightened susceptibility to regulatory element interactions. Furthermore, by correlating target gene expression with ChIP-seq signals, we observed tissue specific expression patterns across species, with notably elevated expression in seeds, roots, and stems, reflecting the functional and structural diversification within Brassicaceae. In summary, our regulatory element centric approach not only expands the genetic toolkit for studying high-level gene regulation but also provides novel mechanistic insights into Brassicaceae biology, with potential implications for other species.