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Abstract Background B-type lamins are critical nuclear envelope proteins that interact with the three-dimensional genomic architecture. However, identifying the direct roles of B-lamins on dynamic genome organization has been challenging as their joint depletion severely impacts cell viability. To overcome this, we engineered mammalian cells to rapidly and completely degrade endogenous B-type lamins using Auxin-inducible degron technology. Results Using live-cell Dual Partial Wave Spectroscopic (Dual-PWS) microscopy, Stochastic Optical Reconstruction Microscopy (STORM), in situ Hi-C, CRISPR-Sirius, and fluorescence in situ hybridization (FISH), we demonstrate that lamin B1 and lamin B2 are critical structural components of the nuclear periphery that create a repressive compartment for peripheral-associated genes. Lamin B1 and lamin B2 depletion minimally alters higher-order chromatin folding but disrupts cell morphology, significantly increases chromatin mobility, redistributes both constitutive and facultative heterochromatin, and induces differential gene expression both within and near lamin-associated domain (LAD) boundaries. Critically, we demonstrate that chromatin territories expand as upregulated genes within LADs radially shift inwards. Our results indicate that the mechanism of action of B-type lamins comes from their role in constraining chromatin motion and spatial positioning of gene-specific loci, heterochromatin, and chromatin domains. Conclusions Our findings suggest that, while B-type lamin degradation does not significantly change genome topology, it has major implications for three-dimensional chromatin conformation at the single-cell level both at the lamina-associated periphery and the non-LAD-associated nuclear interior with concomitant genome-wide transcriptional changes. This raises intriguing questions about the individual and overlapping roles of lamin B1 and lamin B2 in cellular function and disease.

Abstract Background The mutational processes underlying non-coding cancer mutations and their biological significance in tumor evolution are poorly understood. To get better insights into the biological mechanisms of mutational processes in breast cancer, we integrate whole-genome level somatic mutations from breast cancer patients with chromatin states and transcription factor binding events. Results We discover that a large fraction of non-coding somatic mutations in estrogen receptor (ER)-positive breast cancers are confined to ER binding sites. Notably, the highly mutated estrogen receptor binding sites are associated with more frequent chromatin loop contacts and the associated distal genes are expressed at higher level. To elucidate the functional significance of these non-coding mutations, we focus on two of the recurrently mutated estrogen receptor binding sites. Our bioinformatics and biochemical analysis suggest loss of DNA-protein interactions due to the recurrent mutations. Through CRISPR interference, we find that the recurrently mutated regulatory element at the LRRC3C-GSDMA locus impacts the expression of multiple distal genes. Using a CRISPR base editor, we show that the recurrent C→T conversion at the ZNF143 locus results in decreased TF binding, increased chromatin loop formation, and increased expression of multiple distal genes. This single point mutation mediates reduced response to estradiol-induced cell proliferation but increased resistance to tamoxifen-induced growth inhibition. Conclusions Our data suggest that ER binding is associated with localized accumulation of somatic mutations, some of which affect chromatin architecture, distal gene expression, and cellular phenotypes in ER-positive breast cancer.