Duba, I. | Search bioRxiv

bioRxiv2025

Direct measurement of sub-kilobase chromatin structure reveals that linker histone H1 broadly compacts chromatin, with differential impact amongst epigenetic states

Chromatin compaction by linker histone H1 family proteins is a long-standing model for transcriptional repression. However, the biophysical and conformational details of such compaction in situ, at the kilobase- and sub-kilobase length scale relevant to the activity of transcriptional regulatory elements, remain under debate. Rather than inferring such compaction from indirect measurements of features like DNA accessibility, we sought to directly probe sub-kilobase contacts between nearby nucleosomes. We developed an improved version of radiation-induced correlated cleavage with sequencing (RICC-seq), which we term RICC-seq 2.0, and used it in parallel with Micro-C to cross-validate our measurements of chromatin structure in both diverse cell types with different levels of linker histone and different levels of chromatin compaction, as well as a CRISPRi system for pan-H1 depletion. Using this system, we find that chromatin fiber de-compaction upon H1 depletion is global across the genome, reducing the contrast in inter-nucleosome contacts between acetylated chromatin and the rest of the genome. Surprisingly, this does not dramatically change higher-order chromatin organization such as nuclear compartments. Nevertheless, we observe a broad increase in accessibility at tens of thousands of sites and an increase in expression of over a thousand genes, which are enriched in polycomb repressive complex targets. Investigating the local chromatin compaction at upregulated genes as opposed to genes that do not change transcription, we observe that upregulated genes are not specifically de-compacted. Rather, our data support a model in which linker histone globally induces local compaction of nucleosome contacts and an increase in linker lengths, and repression by PRC1/2 is particularly dependent on these local features of chromatin architecture.

genomics↗

bioRxiv2025

CDK4/6 inhibition induces a DNA damage-independent senescence-associated secretory phenotype driven by delayed activation of NF-κB

Cellular senescence consists of regulated cell phenotypes associated with permanent exit from the cell cycle in response to stressors such as genomic instability. The consequences of senescence go beyond individual cells due to the senescence associated secretory phenotype (SASP), which can induce inflammation in neighboring cells. Some cancers respond to CDK4/6 inhibitors (CDK4/6i)--a family of targeted therapies that inhibit proliferation--with a senescence-like phenotype in the absence of DNA damage. We asked how the SASP and the transcriptional regulatory profile triggered by CDK4/6i-driven arrest compares to the canonical NF-{kappa}B-regulated SASP triggered by DNA damage. We profiled the temporal dynamics of transcriptional regulation in response to the CDK4/6i, palbociclib, and the DNA damaging agent, doxorubicin. We found that, although upregulation of NF-{kappa}B driven-SASP genes is shared across both drugs, it is delayed in CDK4/6i. This coincides with slower enhancer activation and epigenetic changes. Interestingly, ATM/ATR inhibition does not affect CDK4/6i-induced NF-{kappa}B nuclear localization, pointing to an alternative mechanism driving NF-{kappa}B activity in the absence of DNA damage. Inhibiting NF-{kappa}B suppresses the expression of shared SASP genes without reversing stable arrest. This points to SASP manipulation as a potential therapeutic strategy, and resolves an ongoing controversy about the nature of cell cycle arrest-driven SASP.

cancer biology↗