Spatio-Temporal Organization of the Super-Enhancer Structure in Cancer Cells

Our research investigates the spatiotemporal organization of nuclear architecture via biomolecular condensation, focusing on the role of liquid–liquid phase separation (LLPS) in gene regulation during cancer progression. We identify phosphatidylinositol 4,5 bisphosphate (PIP2) as a key nuclear lipid forming membrane less condensates with RNA, affecting transcriptional regulators and super-enhancer assembly. PIP2 shows RNA- dependent nuclear localization and interacts with RNA polymerases I and II, suggesting a structural role in transcriptionally active chromatin. Mass spectrometry revealed the RNA-dependent PIP2-associated (RDPA) nuclear proteome, enriched for intrinsically disordered regions and polybasic motifs, supporting LLPS. Among RDPA proteins, BRD4 undergoes PIP2- and RNA-dependent condensation. Using a system for inducible nuclear PIP2 depletion, we demonstrate its role in organizing transcriptional condensates and modulating super-enhancer hubs. These findings establish PIP2-driven phase separation as a key mechanism in nuclear domain organization and cancer-related gene regulation. This project is supported by the project National Institute for Cancer Research (Programme EXCELES, ID Project No. LX22NPO5102) by the European Union – Next Generation EU.