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In this study, we delve into the impact of genotoxic anticancer drug treatment on the chromatin structure of human cells, with a particular focus on the effects of doxorubicin. Using Hi-C, ChIP-seq, and RNA-seq, we explore the changes in chromatin architecture brought about by doxorubicin and ICRF193. Our results indicate that physiologically relevant doses of doxorubicin lead to a local reduction in Hi-C interactions in certain genomic regions that contain active promoters, with changes in chromatin architecture occurring independently of Top2 inhibition, cell cycle arrest, and differential gene expression. Inside the regions with decreased interactions, we detected redistribution of RAD21 around the peaks of H3K27 acetylation. Our study also revealed a common structural pattern in the regions with altered architecture, characterized by two large domains separated from each other. Additionally, doxorubicin was found to increase CTCF binding in H3K27 acetylated regions. Furthermore, we discovered that Top2-dependent chemotherapy causes changes in the distance decay of Hi-C contacts, which are driven by direct and indirect inhibitors. Our proposed model suggests that doxorubicin-induced DSBs cause cohesin redistribution, which leads to increased insulation on actively transcribed TAD boundaries. Our findings underscore the significant impact of genotoxic anticancer treatment on the chromatin structure of the human genome.

The relationship between chromatin organization and gene regulation remains unclear. While disruption of chromatin domains and domain boundaries can lead to misexpression of developmental genes, acute depletion of regulators of genome organization has a relatively small effect on gene expression. It is therefore uncertain whether gene expression and chromatin state drive chromatin organization or whether changes in chromatin organization facilitate cell-type-specific activation of gene expression. Here, using the dorsoventral patterning of the Drosophila melanogaster embryo as a model system, we provide evidence for the independence of chromatin organization and dorsoventral gene expression. We define tissue-specific enhancers and link them to expression patterns using single-cell RNA-seq. Surprisingly, despite tissue-specific chromatin states and gene expression, chromatin organization is largely maintained across tissues. Our results indicate that tissue-specific chromatin conformation is not necessary for tissue-specific gene expression but rather acts as a scaffold facilitating gene expression when enhancers become active.

Background: Precise quantitative and spatiotemporal control of gene expression is necessary to ensure proper cellular differentiation and the maintenance of homeostasis. The relationship between gene expression and the spatial organisation of chromatin is highly complex, interdependent and not completely understood. The development of experimental techniques to interrogate both the higher-order structure of chromatin and the interactions between regulatory elements has recently lead to important insights on how gene expression is controlled. The ability to gain these and future insights is critically dependent on computational tools for the analysis and visualisation of data produced by these techniques., Results and Conclusion: We have developed GenomicInteractions, a freely available R/Bioconductor package designed for processing, analysis and visualisation of data generated from various types of chromosome conformation capture experiments. The package allows the easy annotation and summarisation of large genome-wide datasets at both the level of individual interactions and sets of genomic features, and provides several different methods for interrogating and visualising this type of data. We demonstrate this package's utility by showing example analyses performed on interaction datasets generated using Hi-C and ChIA-PET.

Chromosome conformation capture approaches have shown that interphase chromatin is partitioned into spatially segregated Mb-sized compartments and sub-Mb-sized topological domains. This compartmentalization is thought to facilitate the matching of genes and regulatory elements, but its precise function and mechanistic basis remain unknown. Cohesin controls chromosome topology to enable DNA repair and chromosome segregation in cycling cells. In addition, cohesin associates with active enhancers and promoters and with CTCF to form long-range interactions important for gene regulation. Although these findings suggest an important role for cohesin in genome organization, this role has not been assessed on a global scale. Unexpectedly, we find that architectural compartments are maintained in noncycling mouse thymocytes after genetic depletion of cohesin in vivo. Cohesin was, however, required for specific long-range interactions within compartments where cohesin-regulated genes reside. Cohesin depletion diminished interactions between cohesin-bound sites, whereas alternative interactions between chromatin features associated with transcriptional activation and repression became more prominent, with corresponding changes in gene expression. Our findings indicate that cohesin-mediated long-range interactions facilitate discrete gene expression states within preexisting chromosomal compartments.