The loss of heterochromatin is associated with multiscale three-dimensional genome reorganization and aberrant transcription during cellular senescence.

Heterochromatin remodeling is critical for various cell processes. In particular, the "loss of heterochromatin" phenotype in cellular senescence is associated with the process of aging and age-related disorders. Although biological processes of senescent cells, including senescence-associated heterochromatin foci (SAHF) formation, chromosome compaction, and redistribution of key proteins, have been closely associated with high-order chromatin structure, the relationship between the high-order chromatin reorganization and the loss of heterochromatin phenotype during senescence has not been fully understood. By using senescent and deep senescent fibroblasts induced by DNA damage harboring the "loss of heterochromatin" phenotype, we observed progressive 3D reorganization of heterochromatin during senescence. Facultative and constitutive heterochromatin marked by H3K27me3 and H3K9me3, respectively, show different alterations. Facultative heterochromatin tends to switch from the repressive B-compartment to the active A-compartment, whereas constitutive heterochromatin shows no significant changes at the compartment level but enhanced interactions between themselves. Both types of heterochromatin show increased chromatin accessibility and gene expression leakage during senescence. Furthermore, increased chromatin accessibility in potential CTCF binding sites accompanies the establishment of novel loops in constitutive heterochromatin. Finally, we also observed aberrant expression of repetitive elements, including LTR (long terminal repeat) and satellite classes. Overall, facultative and constitutive heterochromatin show both similar and distinct multiscale alterations in the 3D map, chromatin accessibility, and gene expression leakage. This study provides an epigenomic map of heterochromatin reorganization during senescence.