Under a variety of stressful conditions, cells develop small compartments that are transient. These compartments are termed dynamic intracellular bodies. Little is known about the molecular interactions that form these bodies. Many of these bodies contain RNA-protein complexes that could protect cellular RNAs during cell stress. The goal of this research is to illuminate the interactions between proteins and of proteins with RNAs that lead to the formation of these bodies. This will be achieved using a variety of analyses using structural tools such as X-Ray crystallography and electron microscopy backed up by biochemical studies. While the research is of a fundamental nature, some of the proteins that are involved in the formation of these dynamic intracellular bodies are also associated in aggregated form with Amyotrophic Lateral Sclerosis (ALS), so the structural studies may provide the additional benefit of understanding the connection of disease. A part of the research is also focused on developing novel methods for structure determination, such as micro-electron diffraction. The broader societal significance involves the recruitment into scientific careers of students and postdoctoral researchers from underrepresented groups. Additionally, through the UCLA Center for Global Mentoring, the project aims to spread the American system of scientific mentoring to countries with developing scientific infrastructures.
To fill the void of knowledge about the molecular interactions in dynamic intracellular bodies, also known as membraneless organelles, the project will discover the structural basis of a newly recognized form of protein-protein and protein-RNA interaction. In Aim 1, it has been discovered that the atomic structures of the adhesive segments of reversible amyloid-like fibrils (RALFs) formed by the low complexity sequences have both similarities and differences with disease-related amyloid. The project will explore the full variety of interactions and assembly states found in membraneless organelles. In other words, the human reversible amylome will be mapped. In Aim 2, it is hypothesized that RNA binding can strongly influence membraneless organelle and reversible amyloid formation through several parallel mechanisms. These mechanisms will be tested using biochemical and cellular assays. The contributions of RALFs and protein-RNA interactions will also be compared. Both Aims will deepen understanding of cellular organization and function, particularly mechanisms for sequestration of proteins and RNAs without the aid of membrane encapsulation. Part of the research involves determination of structures from miniscule amyloid-like crystals. The frontier method of micro-electron diffraction must be further developed.
