Dynamic reorganization of three-dimensional genome architecture during Populus diversification.

Advancements in sequencing technology enable the investigation of genomic information within its three-dimensional (3D) spatial structure, offering new insights into genome organization and function. However, the role of higher-order chromatin architecture in species divergence and speciation remains largely unexplored. Here we constructed 3D genome maps and performed comparative analyses across 11 phylogenetically diverse Populus species, revealing that evolutionary changes in 3D chromosomal architecture are closely associated with genomic differentiation. While chromatin compartments (A/B) are generally conserved across species, topologically associated domains exhibit substantial divergence. Integrating multi-omics data, we observed that genes located in dynamic 3D chromatin regions display distinct patterns of sequence conservation, gene expression and epigenetic modifications, emphasizing the interplay among spatial chromatin reorganization, transcriptional regulation and epigenetics during genome evolution. Structural variants are shown to play a key role in shaping interspecific 3D genome diversity. Notably, a ~76-bp insertion/deletion in HSFA2 3' UTR within divergent topologically associated domain boundaries modulates divergent heat-stress responses across species, underscoring the potential functional significance of 3D genome changes in adaptive evolution. Together, this study highlights the necessity of integrating 3D chromatin organization with genomic and epigenomic variation to dissect the molecular mechanisms underlying species divergence and ecological adaptation.