Collaborative Research: Transcriptome-wide visualization and function of RNA tertiary structure

RNA serves as the central messenger that carries genetic instructions in all living cells. RNA molecules fold into complex three-dimensional shapes that control how genes are turned on and off. While scientists have identified many RNAs with complex structures – and critical functions – in bacteria and viruses, very few have been discovered in human cells, leaving a major gap in understanding how genes are regulated in humans. This project addresses this knowledge gap by using newly developed technologies to systematically discover, characterize, and visualize RNA structures throughout human cells. The research team has already identified hundreds of previously unknown RNA structures in human cells and developed innovative methods to determine their precise three-dimensional shapes. This work serves the national interest by, first, advancing fundamental knowledge of gene regulation with applications in developing new RNA-targeted medicines. Second, the three-dimensional RNA structures solved will support artificial intelligence efforts to predict RNA structures, supporting breakthroughs analogous to those achieved in protein folding. Third, the project will train undergraduate researchers in collaborative team science, preparing the next generation of biotechnology innovators and strengthening the US scientific workforce in an area of strategic importance. This project will systematically identify, functionally characterize, and solve high-resolution structures of RNA tertiary motifs discovered using an innovative chemical probing technology that selectively detects three-dimensional RNA folds. The research goals are to: (1) investigate functional roles of RNA tertiary structures in transcription factor genes; (2) evaluate regulatory impacts of tertiary structures in 5'-untranslated regions using massively parallel reporter assays; (3) establish an efficient pipeline for solving RNA tertiary structures using an innovative cryo-electron microscopy scaffold, with the aim of resolving multiple novel RNA 3D structures to better than 3.0 Å resolution; and (4) create an undergraduate collaboratory focused on mechanistic understanding and structure determination of newly identified RNA tertiary structures. The project integrates iterative cycles of transcriptome-wide discovery, functional validation, and high-resolution structural imaging. By creating a comprehensive understanding of complex RNA structures and developing streamlined cryo-EM strategies, this work will address critical limitations in current machine learning approaches for RNA structure prediction and will establish foundational knowledge needed for designing RNA-targeted therapeutics, advancing both artificial intelligence and biotechnology applications in the biomedical sciences. 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. NSF Award ID: 2550997 | Program: 01002627DB NSF RESEARCH & RELATED ACTIVIT | Principal Investigator: Navtej Toor | Institution: University of California-San Diego, LA JOLLA, CA | Award Amount: $800,000 View on NSF Award Search: https://www.nsf.gov/awardsearch/show-award/?AWD_ID=2550997 View on Research.gov: https://www.research.gov/awardapi-service/v1/awards/2550997.html