DecipheringPhase-Separated Mitochondrial RNA Granulesunder Stress Conditions with the Mitoribosome-Targeting Small Molecule

RNA granules are liquid–liquid-phase-separated condensates comprising RNA and proteins. Despite growing insights into their biological functions, studies have predominantly relied on biological methodologies lacking adequate chemical tools. Here, we introduce ICP-CHARGINGS, a concept for efficiently identifying chemical probes to characterize RNA granules of interest among nucleic acid-targeting agents. Focusing on mitochondrial RNA granules (MRGs), whose functions remain elusive, we developed a methodology within this framework and identified <b>NATA</b>, a new fluorescent molecule that, following mechanistic studies, was found to bind to the mitoribosome, enabling MRG labeling and recognition. Using <b>NATA</b> to reveal the potential buffering roles of MRGs, we demonstrated a close correlation between MRG maintenance and assembly and cellular survival and proliferation under cold shock and hypoxic stress. Overall, the introduction and implementation of the ICP-CHARGINGS strategy provide a specialized chemical tool for advancing our comprehension of MRG biology and establish a paradigm for elucidating RNA structures within RNA granules that can be targeted by small molecules, paving the way for developing tailored chemical probes for diverse RNA granules in future research.