Exclusion of RNA-binding domains from G-quadruplex condensates by G-quadruplex ligands.

Biomolecular liquid-liquid phase separation, which forms droplets in living cells, plays a crucial role in the regulation of gene expression. Dysfunction of liquid-liquid phase separation leads to aberrant aggregates that sequester RNA-binding proteins, thereby impairing their functions, which potentially leads to the onset of neurodegenerative diseases. Preventing and reversing the sequestration of RNA-binding proteins from droplets represent a promising therapeutic approach. Noteworthily, the G-quadruplex, a non-canonical secondary structure of nucleic acids formed by a guanine-rich sequence, is an essential structural motif that triggers liquid-liquid phase separation with a partner protein. Thus, a G-quadruplex ligand, which selectively binds and stabilizes the G-quadruplex, can be promising for controlling liquid-liquid phase separation of G-quadruplexes and partner proteins. In this study, we investigated the effects of G-quadruplex ligands on the liquid-liquid phase separation of RNA G-quadruplexes and RGG domain-derived cationic peptides. It was found that G-quadruplex ligands formed aggregates with the target G-quadruplexes and excluded the G-quadruplex-binding peptides from these aggregates. Moreover, structure-selective G-quadruplex ligands induced aggregates only with the G-quadruplex but not with other secondary structures. These findings demonstrate for the first time that the structure-selectivity of G-quadruplex ligands plays a key role in modulating condensates.