Intracellular communication through designer membraneless organelles

Back in the 1950s, the membranous nature of optically prominent cytoplasmic compartments such as the nucleus and mitochondria was discovered, each housing a distinct set of proteins enabling specialized functions. However, just over the last decade, we have begun to understand that cells also contain numerous and dynamic membraneless organelles, also known as biomolecular condensates, that form and dissolve as needed. De-mixing of multivalent biomolecules via the process of liquid-liquid phase separation is crucial to their formation, and intrinsically disordered regions (IDRs) of proteins are found to play an active role in many of these condensates.1 We would like to utilize IDRs to compartmentalize synthetic cells, bottom-up mimics of living cells, and address intracellular communication through designer phase-separating membraneless organelles (MOs) in cell-like vesicles using on-chip microfluidic techniques, advanced microscopy, and molecular cloning. The idea is to generate multiple, functional, condensate-based compartments within synthetic cells to drive independent enzymatic reactions. The methodology involves making use of well-characterized pH-responsive elastin-like polypeptides (PREs) consisting of repetitive pentapeptide sequences2 and designing a family of their surfactant counterpart to stabilize the surface of corresponding MOs, preventing mixing. Furthermore, we will make use of our microfluidic expertise to make a variety of vesicles, such as double emulsions3 and liposomes4 in a controlled manner, making it an effective setup to study the behavior of the PREs inside cell-mimicking confinements.

References:
1. Hirose, Tetsuro, et al. "A guide to membraneless organelles and their various roles in gene regulation." Nature Reviews Molecular Cell
2. de Haas, Robbert J., et al. "pH-responsive elastin-like polypeptide designer condensates." ACS applied materials & interfaces
3. Chen, Chang, et al. "Elastin-like polypeptide coacervates as reversibly triggerable compartments for synthetic cells." Communications Chemistry
4. Deshpande, Siddharth, et al. "Octanol-assisted liposome assembly on chip." Nature communications