Background

The nucleolus is the cellular hub for ribosome biogenesis, where RNA polymerase I (Pol I) transcribes rRNA genes. Because many cancers depend on elevated ribosome production, Pol I inhibitors such as CX-5461 have attracted attention as anti-cancer agents. However, beyond blocking rRNA synthesis, it has remained unclear how Pol I inhibition reshapes nuclear organization and chromatin regulation.

NPM1 is a major nucleolar protein involved in ribosome biogenesis, RNA binding, chromatin regulation, and nucleocytoplasmic shuttling. NPM1 mutations are highly relevant in leukemia, and reduced NPM1 levels sensitize cells to CX-5461. This suggested that NPM1 might connect nucleolar stress to broader epigenetic and 3D genome changes. 

Question

The study asked: How does inhibition of RNA Pol I alter nucleolar structure and NPM1 localization? Does NPM1 relocalization drive epigenetic remodeling of chromatin?  Can nucleolar disruption reorganize higher-order genome architecture, including NADs, LADs, compartments, and chromatin loops?

Main FindingsRNA Pol I inhibition rapidly redistributes NPM1

After CX-5461 treatment, NPM1 moved away from the nucleolus and accumulated near the nuclear lamina. This occurred together with nuclear structural abnormalities and lamina detachment. RNA digestion experiments showed that NPM1 localization depends strongly on RNA interactions, especially rRNA.

NPM1 protects active chromatin states

Under normal conditions, NPM1 was associated with active chromatin marks such as H3K9ac. When Pol I was inhibited or NPM1 was depleted, HDAC1 loaded onto chromatin, removing H3K9 acetylation. This exposed H3K9 for methylation by SUV39H1, increasing the repressive mark H3K9me3. Thus, NPM1 functions as a gatekeeper that preserves active chromatin and prevents heterochromatin spreading.

Chromatin becomes less accessible and more methylated

Genome-wide open chromatin was reduced after CX-5461 treatment, especially at promoters and enhancers. These regions gained H3K9me3 and DNA methylation, with increased recruitment of DNMT1. This indicates a broad shift toward transcriptional repression.

Nuclear domains are reorganized

Nucleolus-associated domains (NADs) were remodeled and increasingly overlapped with lamin-associated domains (LADs). Facultative heterochromatin-like regions decreased, while constitutive heterochromatin expanded. This suggests chromatin territories moved from nucleolar association toward the nuclear periphery.

3D genome structure is altered

Hi-C analysis showed expansion of B compartments (inactive chromatin), loss of A compartments (active chromatin), enlarged TADs, and loss of many chromatin loops, especially enhancer-promoter interactions. Therefore, nucleolar transcription status influences genome topology. 

Key methods

This study identified RNAs bound by NPM1 by RIP-seq and mapped genome-wide binding of NPM1, Pol I, lamin B2, histone marks by NEED-seq.

My question: Why CX-5461 treatment induces lamina detachment?

article linker: NPM1 mislocalization mediated by RNA Pol I inhibition alters chromatin landscape - PubMed

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