Effects of topological domain disruption on transcriptional regulation are chromatin context dependent

ABSTRACT: The three-dimensional (3D) chromatin organization within the nucleus is essential for regulating gene expression by facilitating interactions between cis-regulatory elements while insulating domains to restrict non-specific interactions. CTCF and cohesin, key architectural proteins, form topologically associated domains (TADs) that play a significant role in this spatial organization. While previous studies have demonstrated that TAD boundaries function to insulate genes from inappropriate interactions with distal enhancers or silencers, the functional impact of these boundaries on gene expression remains highly complex and context dependent. Here, we investigate the topological organization of the Notch1 locus, a gene involved in cell fate determination and cancer, to understand the regulatory impact of TAD boundaries on gene expression. We identified a highly conserved topological domain surrounding Notch1, with CTCF and cohesin binding at both 5' and 3' boundaries. Deletion of these boundary elements via CRISPR-Cas9 led to a significant increase in Notch1 expression, as well as enhanced expression of neighboring non-coding RNAs (Nalt1 and Linc01451) within the domain. Disruption of these boundaries also resulted in altered cell proliferation and colony formation. These effects correlated with altered enhancer-promoter interactions within the domain, especially involving a key enhancer located within the Linc01451 gene. Cross-species analyses revealed that the Notch1 domain and its boundary organization are conserved in mice, where perturbations similarly influenced Notch1 expression and cellular phenotypes in mouse embryonic stem cells (mESCs) but had minimal effects in murine erythroleukemia (MEL) cells. Together, these findings suggest that CTCF-dependent topological boundaries play a critical role in insulating the Notch1 locus and maintaining precise expression levels in a chromatin context- dependent manner, highlighting a regulatory mechanism that may extend to other lineage-specific and disease-relevant genes.

ORGANISM(S): Homo sapiens

PROVIDER: GSE296362 | GEO | 2026/03/04

REPOSITORIES: GEO