Abstract Biomolecular condensates require suitable material properties to properly carry out their function. Yet, modulators that affect the material properties of condensates have remained largely unexplored. Here we apply Differential Dynamic Microscopy (DDM) to probe the material properties of an in vitro model of processing bodies (P-bodies) consisting of condensates formed by the DEAD-box ATPase Dhh1 in the presence of ATP and RNA. DDM allows us to measure the viscosity of liquid droplets and to distinguish between liquid-like and gel- or glass-like condensates. By applying this single-droplet technique we show that condensates within the same population exhibit a distribution of material properties, which can be drastically affected by several modulators such as the low-complexity domains (LCDs) of the protein, the protein/RNA ratio, the type of RNA as well as the enzymatic activity. In particular, we show that structured RNA leads to a large fraction of dynamically arrested condensates with respect to unstructured polyuridylic acid (polyU), emphasizing the role of RNA structure in condensate dynamics. We further demonstrate that the ageing of the condensates and the formation of gel or glass-like structures is reduced by promoting the enzymatic ATPase activity of Dhh1 and the rate of droplet formation and dissolution. Our study shows that not only the reversible formation and dissolution of condensates but also their material properties are regulated on several levels, and that biochemical activity and material turnover can be important to maintain fluid-like properties over time.

Dynamic arrest and aging of biomolecular condensates are regulated by low-complexity domains, RNA and biochemical activity