RNA functions as the central conduit for information in living systems. RNA molecules encode genetic information and also serve as critical component in the catalytic and regulatory machinery in cells. In one example, the ribosome is a finely tuned machine that translates the information coded in messenger RNA into proteins. The ribosome contains two large RNA molecules and many proteins and the function of this essential cellular machine is governed by complex structural and dynamic interactions among its protein and RNA components. In a second example, RNA viruses represent perhaps the most successful replicating entities in our biosphere. Almost every step in RNA virus replication involves highly evolved interactions between their RNA genome and viral and cellular proteins. Extraordinary progress in understanding the ribosome and RNA viruses has been made based on genetic, biochemical, and high-resolution structure approaches. However, the impact of the cellular environment on RNA function remains poorly understood. The overarching vision of this project is to make the SHAPE (selective 2'-hydroxyl acylation analyzed by primer extension) RNA structure analysis technology a robust tool for analysis of RNA structure in cells and inside viral particles. The specific research goals over five years of the project are to use SHAPE to establish the structure of E. coli ribosomal RNA during protein synthesis in healthy, growing cells and to determine the RNA genome structure of an icosahedral plus-strand virus to reveal broad principles of viral RNA genome packaging.
Broader Impact Accurate and efficient technology for analyzing RNA structure inside cells has the potential to transform many fields in the biological sciences. When fully developed, SHAPE technology will allow synthesis of information obtained from high-resolution structural approaches in highly purified systems with function and fitness studies in cells. SHAPE technology is also sufficiently mature to serve as the basis for a completely undergraduate-driven research endeavor, the Undergraduate Transcriptome Project (the UTP). Undergraduates will, independently, carry out the research on viral RNA genome packaging. The UTP is designed to help undergraduates explore their potential for independent creativity, to fuel their passion for science, to promote high-level mentoring by graduate students, and to be a model for engaging undergraduates in a research university.