IIBR RoL: Applying innovative structural tools to highlight RNA's structural dynamics as RNA-protein complexes self-assemble

Cornell University, Ithaca NY

Investigators

Abstract

Life functions are supported by the interaction of large biological molecules, including proteins and RNA. Although the shapes or structures of some of these complexes have been visualized, very little is known about their dynamics of assembly. How do these differing components interact to create complexes whose function exceeds that of either alone? This is a challenging problem because of the intertwined motions of the components. Through this project, new tools will be applied to highlight the motions of just one macromolecular species during the assembly of a complex. Although both proteins and RNA are present, only the RNA is visible. Students will design, build and apply the new technology to study the self-assembly of a small plant virus, an intricate structure consisting of a protein shell surrounding a nucleic acid core. These studies will allow us to more clearly define the rules that govern the joining of nucleic acids and proteins into biological complexes. Many important biological assembles are comprised of distinct macromolecular components, such as proteins in conjunction with the nucleic acids. Despite their importance, less than 2% of the static structures in the protein data bank reflect RNA-protein complexes. Even less is known about the dynamic interactions between RNA and protein components. Although much effort has been expended in watching one type macromolecule self-assemble, e.g. protein folding, it is much more difficult to interpret dynamic structural information for multicomponent machines because multiple species are present. The inability to separate the signal from the distinct components precludes a simple interpretation. This project will exploit and innovate existing experimental infrastructure to create tools that highlight the signal of only one species on the background of the other, vastly simplifying measurements of complexes' dynamics. Mixers that rapidly combine the protein and RNA precursors will be constructed and will allow experimenters to track the changing structures of only the RNA component during complex assembly. A small plant virus will serve as a model system for these studies. The development and application of two complementary techniques (small angle x-ray scattering and single molecule fluorescence) will offer a unique perspective on this self-assembly problem. Once established, these methods can be applied other RNA-protein complexes, gathering data to elucidate the rules governing their assembly. Project results, sorted by topic, will be posted at: https://pollack.research.engineering.cornell.edu/all_publications/. This project is jointly funded by the Division of Biological Infrastructure program for Infrastructure Innovation for Biological Research, the Molecular Biophysics and Genetic Mechanism Clusters in the Division of Molecular and Cellular Biosciences, and the Rules of Life initiative in the Emerging Frontiers office of the Biology Directorate. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.