Targeted environment-sensitive fluorescent probes for biomolecular condensates – PROBIC

Biomolecular condensates (BCs) are microscopic intracellular fluidic bodies that are formed by certain proteins and RNAs in the process known as liquid-liquid phase separation (LLPS). Biomolecular condensates play many roles in normal cell physiology and homeostasis, whereas misregulation of LLPS has been linked to several pathological states, such as neurodegenerative diseases. The material state of BCs can vary in a broad range, from fluidic droplets to gel-like and solid-like aggregates, depending on the molecular composition of the condensed phase and environmental conditions. Probing viscoelastic properties of the condensates is essential for deciphering their biological roles because these properties control the internal mobility and exchange rates of the condensate-embedded molecules, and therefore can modulate the biological effects. Several microrheology methods have been used for this purpose, but they are often destructive, time-consuming, and yield droplet- and ensemble-averaged information. Here I propose a new way for probing viscoelastic states of BCs using targeted fluorescent probes. Environment-sensitive fluorophores covalently tethered to biomolecules undergoing LLPS will enable fast, non-destructive measurements with sub-droplet resolutions, a level of precision that has not been possible yet. The project is highly multidisciplinary, combining rational molecular design, chemical synthesis, spectroscopic characterization, bioconjugation, and fluorescence imaging. The new fluorescent probes proposed herein would be of interest for a range of specialists in many areas of basic research, such as biochemistry, biophysics, cell and synthetic biology.