Specific protein-RNA interactions are mostly preserved in biomolecular condensates

• Published in: Science advances Vol. 10; no. 10; p. eadm7435
• Main Authors: de Vries, Tebbe, Novakovic, Mihajlo, Ni, Yinan, Smok, Izabela, Inghelram, Clara, Bikaki, Maria, Sarnowski, Chris P, Han, Yaning, Emmanouilidis, Leonidas, Padroni, Giacomo, Leitner, Alexander, Allain, Frédéric H-T
• Format: Journal Article
• Language: English
• Published: United States American Association for the Advancement of Science 08.03.2024
• Subjects: Biochemistry; Biomedicine and Life Sciences; Biomolecular Condensates; Life Sciences; Phase Separation; Preservation, Biological; Ribonucleoproteins; RNA; SciAdv r-articles
• ISSN: 2375-2548; 2375-2548
• DOI: 10.1126/sciadv.adm7435

Many biomolecular condensates are enriched in and depend on RNAs and RNA binding proteins (RBPs). So far, only a few studies have addressed the characterization of the intermolecular interactions responsible for liquid-liquid phase separation (LLPS) and the impact of condensation on RBPs and RNAs. Here, we present an approach to study protein-RNA interactions inside biomolecular condensates by applying cross-linking of isotope labeled RNA and tandem mass spectrometry to phase-separating systems (LLPS-CLIR-MS). LLPS-CLIR-MS enables the characterization of intermolecular interactions present within biomolecular condensates at residue-specific resolution and allows a comparison with the same complexes in the dispersed phase. We observe that sequence-specific RBP-RNA interactions present in the dispersed phase are generally maintained inside condensates. In addition, LLPS-CLIR-MS identifies structural alterations at the protein-RNA interfaces, including additional unspecific contacts in the condensed phase. Our approach offers a procedure to derive structural information of protein-RNA complexes within biomolecular condensates that could be critical for integrative structural modeling of ribonucleoproteins (RNPs) in this form.