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Studying ligand-biomacromolecule interactions provides opportunities for creating new compounds that can efficiently regulate specific biological processes. Ribonucleic acid (RNA) molecules have become attractive drug targets since the discovery of their roles in modulating gene expression, while only a limited number of studies have investigated interactions between ligands and functional RNA molecules, especially those based on nanotechnology. DNA-protected silver nanoclusters (AgNCs) were used to investigate ligand-RNA interactions for the first time in this study. The anthracycline anticancer drug mitoxantrone (MTX) was found to quench the fluorescence of AgNCs. After adding human immunodeficiency virus trans-activation responsive region (TAR) RNA or Rev-response element (RRE) RNA to AgNCs-MTX mixtures, the fluorescence of the AgNCs recovered due to interactions between MTX with RNAs. The binding constants and number of binding sites of MTX to TAR and RRE RNA were determined through theoretical calculations. MTX-RNA interactions were further confirmed in fluorescence polarization and mass spectrometry experiments. The mechanism of MTX-based fluorescence quenching of the AgNCs was also explored. This study provides a new strategy for ligand-RNA binding interaction assay.

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