Single molecule probing of disordered RNA binding proteins

Liquid-liquid phase separation of intrinsically disordered proteins is known to underlie diverse pathologies such as neurodegeneration, cancer, and aging. The nucleation step of condensate formation is of critical importance for understanding how healthy and disease-associated condensates differ. Here, we describe four orthogonal single-molecule techniques that enable molecular tracking of the RNA-protein interaction, RNA-induced oligomerization, and kinetics of nucleation. These approaches allow researchers to directly interrogate the initial steps of liquid-liquid phase separation. For complete details on the use and execution of this profile, please refer to Niaki et al. (2020), Rhine et al. (2020), and Rhine et al. (2022).