Cells can condensate some of their intracellular components into compartments known as membraneless-organelles (also called liquid droplets or protein condensates). Among them are nucleoli, Cajal bodies, P-bodies, and stress granules, all existing as liquid protein-rich droplets within the cell and arising from the condensation of cellular material in a process termed liquid-liquid phase separation. Although termed “membraneless”, these droplets can come into contact, wet, and reshape membranous compartments. Whilst there are some reports exploring membrane remodelling by liquid polymer-rich droplets, this is still an underexplored field, and the molecular origin of membrane reshaping remains unresolved. Here, we use giant unilamellar vesicles (GUVs) either encapsulating aqueous two-phase systems (ATPS) or in contact with soy glycinin condensates to explore the molecular interactions with the membrane when liquid-liquid phase separation takes place near the bilayer. We use a microfluidic setup to have a better control of individual vesicles and the external milieu, together with two-photon excitation fluorescence microscopy to exploit the spectroscopic properties of LAURDAN, an environmentally sensitive fluorescent probe. Hyperspectral imaging and fluorescence lifetime imaging (FLIM) combined with a multidimensional phasor analysis, provides us with spatial and time resolved information on these systems. The phasor plot analysis allows us to evaluate the changes that occur when the polymers/protein condensates interact and wet the membrane, enabling us to identify regions of different hydration properties within the membranes. Altogether, this approach provides new information on the relationship between membrane wetting, hydration, and spontaneous curvature generation driven by protein/polymer liquid-liquid phase separation.

Liquid-liquid phase separation modulates membrane structure and hydration

ACS Spring 2022

The official archive and repository for all ACS Meetings & Expositions abstracts and select posters or presentations.