CAREER: Structural and mechanistic studies of RNA-mediated enteroviral genome replication

Enteroviruses have a single-strand (+)-sense RNA genome containing complex RNA elements that direct genome replication by recruiting viral and host proteins to form ribonucleoprotein (RNP) complexes required for viral replication. Knowledge of the 3D structures of these RNA elements and corresponding RNPs is essential for understanding fundamental virological processes. The project will investigate the structural and mechanistic underpinnings of RNA-mediated enteroviral genome replication and determine how RNA structures exploit viral and host proteins to promote replication during viral infections. The findings can inform future therapeutic or preventive strategies to combat pathogens that cause human diseases and global pandemics. The project will also provide opportunities for RNA-centric research to high school, undergraduate, and graduate students, encouraging them to pursue higher studies and careers in RNA biochemistry, structural biology, or related STEM fields. The research will utilize antigen-binding antibody fragments (Fabs) as chaperones for determining the crystal structures of enteroviral replication-linked RNAs and RNPs and use biochemistry and molecular biology tools to elucidate RNA-RNA, RNA-protein, and protein-protein interactions. The objectives are to 1) determine the crystal structures of 5′ cis-acting replication elements (CREs) from seven major enteroviral genotypes (enterovirus A-C and rhinovirus A-D), 2) elucidate the interactions of these 5′CREs with viral 3C, 3CD fusion, and human PCBP proteins, and 3) investigate the crystal structures of 3′CREs and their RNPs formed within the 3′-end of the (-)-sense RNA replication intermediate. The outcomes are expected to reveal structure-function relationships that will provide new insights into the replication mechanism of this major class of viral pathogens. This project is jointly funded by the Genetic Mechanisms and Molecular Biophysics programs of the Molecular and Cellular Biosciences Division in the Biological Sciences 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.