<p>Higher-order chromatin organization plays an important role in gene regulation; however, its tissue-specific features and regulatory relationships remain to be systematically characterized. In this study, we constructed high-resolution three-dimensional (3D) genome maps of pig liver and muscle by integrating Hi-C data with ATAC-seq, ChIP-seq, and RNA-seq datasets. Tissue-differential 3D structures, including compartments, topologically associating domains (TADs), and chromatin loops, exhibited limited spatial overlap and did not show a strictly coordinated hierarchical cascade among these structural layers. We systematically characterized the epigenetic modification features associated with different hierarchical levels of 3D chromatin structure and further analyzed their associations with gene expression. At a subset of genomic loci, compartment status, TAD organization, and enhancer-promoter interactions exhibited coordinated spatial patterns, suggesting that multi-layered chromatin organization might be associated with gene regulation. Further analyses showed that tissue-specific loop anchors tended to be enriched for tissue-specific transcription factors (TFs), and their presence was associated with stronger chromatin interaction signals and higher gene expression levels compared with anchors lacking such binding sites. Our results also supported the possibility that the formation of some tissue-specific chromatin loops was associated with chromatin accessibility, active histone modifications, and coordinated TF enrichment, rather than being entirely dependent on CTCF enrichment. Overall, these results provided a useful reference for understanding tissue-specific gene regulation in mammals and provided a foundational data resource for future functional genomics studies in livestock.</p>