Sequence determinants of membraneless organelle rheology

PROJECT SUMMARY/ABSTRACT The long-term vision of my lab is to elucidate biophysical principles of mesoscale assembly in biology and to harness those discoveries for biomedical and biotechnological advances. In that vein, the lab’s current focus is on elucidating principles underlying intracellular phase separation. Phase separation is a fundamental process that cells use to organize their myriad biomolecules into functional compartments, giving rise to membraneless organelles such as stress granules, germ granules, and the nucleolus, with important roles in gene regulation, stress response, and many additional essential functions. Conversely, aberrant phase transitions are associated with neurodegenerative and other diseases. The overall goal of this MIRA proposal is to establish how biomolecular sequence and composition determine the material properties of membraneless organelles, and how material properties contribute to biological function and misfunction. We will dissect the sequence features and molecular interactions that determine the rheology (liquid-like, solid-like, or a combination) of biomolecular condensates formed by phase separation of intrinsically disordered proteins. We will accomplish this using protein engineering, mutagenesis, microscopy, a suite of biophysical techniques, and nanotechnology, including the development of a toolbox of nanoparticle probes for measuring membraneless organelle rheology. We will then investigate how membraneless organelle rheology underlies biological function, focusing on germ granules, and we will investigate how perturbations to membraneless organelle rheology perturb biological function. Finally, we will examine whether certain sequence features are particularly prone to aberrant liquid-solid phase transitions and toxicity, indicating possible origins of pathology and targets for therapies. Together, this work seeks to provide a strong foundation for understanding the links between molecular sequence, rheology, and function of membraneless organelles in health and disease.