Using single-cell genomics to explore transcriptional divergence and cis -regulatory dynamics of duplicated genes

Gene duplication is a major source of evolutionary innovation, enabling the emergence of novel expression patterns and functions. Leveraging single-cell genomics, we investigated the transcriptional dynamics and cis -regulatory evolution of duplicated genes in cultivated soybean ( Glycine max ), a species that has undergone two rounds of whole-genome duplication. Our analysis revealed extensive transcriptional divergence among these duplicated gene pairs, including dosage balanced, paralogue dominance, divergence, specialization, and widespread mono-expressed pattern. These expression divergences are correlated with sequence variation in their associated accessible chromatin regions (ACRs), where cis -regulatory elements reside. Moreover, different duplication mechanisms, including whole-genome duplication and small-scale duplication, likely give rise to distinct types of cis- regulatory variants, leading to varying effects on transcriptional divergence. To further explore the evolution of cis -regulatory elements and their impact on gene expression, we focused on gene sets in which duplicated genes are derived from two rounds of whole-genome duplication and share a common ancestral gene, as cis -regulatory elements are typically co-duplicated with genes during whole-genome duplication. Most ACRs were retained after duplication whereas a subset likely arose de novo . Both mutations in duplicated elements and the emergence of de novo evolved elements contribute to widespread expression divergence within these gene sets. We also characterized the evolution of cell-type-specific expression and cell-type-specific ACRs among these sets. Collectively, our findings highlight the important role of cis- regulatory evolution in shaping transcriptional divergence and facilitating the retention of duplicated genes, offering a high-resolution view through the lens of single-cell genomics.