Enabling High-Throughput RNA Structural Biology

RNAs play diverse and critical roles across the cell by folding into intricate structures that orchestrate molecular interactions at key times and places. We know that RNA folding creates strong helices that can prevent protein binding and can regulate RNA degradation and protein synthesis. RNA folding also creates binding pockets for ligands, allowing RNAs to act as molecular switches that regulate cascades of biochemical processes, such as one-carbon metabolism. To fully understand the mechanisms of RNA function in biological systems, we now must grapple with the challenge of elucidating these structures, or in other words solve the cellular RNA folding problem.  

Two central questions of our research are, how do RNAs fold inside the cell? And, how do these folds mediate RNA function? To address this, we develop high-throughput techniques that uncover the detailed mechanisms of how RNAs fold and function across the cell. This technology platform, SHAPE-Seq, allows acquisition of nucleotide-resolution structural information for thousands of RNAs in a single experiment, using chemical probing and next-generation sequencing to chemically “image” RNA structures. We have expanded the technology to create new paradigms for study of RNA folding and function, including in-cell SHAPE-Seq. We are now interested in transforming high-throughput probing of RNA structure from a technique that measures RNA structures to one that measures RNA structural dynamics. Such a technique may be able to more robustly uncover higher-order structural features of RNAs such as tertiary structures and non-canonical interactions, often the centerpieces of ligand binding and catalytic and regulatory function of RNAs.   

Featured Publications: 
1. High-throughput determination of RNA structures.

E. J. StrobelA. M Yu, J. B. LucksNature Reviews Genetics. (2018).
Links: JournalPDF

2. Simultaneous characterization of cellular RNA structure and function with in-cell SHAPE-Seq.
K. E. Watters, Timothy R. Abbott, J. B. LucksNucleic Acids Research. (2015).
Links: JournalPDF

3. Characterizing RNA structures in vitro and in vivo with selective 2′-hydroxyl acylation analyzed by primer extension sequencing (SHAPE-Seq).
K. E. Watters, A. M. Yu, E. J. Strobel, A. H. Settle, J. B. LucksMethods. (2016).
Links: JournalPDFBioRxiv (Open Access)

4. SHAPE-Seq 2.0: Systematic optimization and extension of high-throughput chemical probing of RNA secondary structure with next-generation sequencing.
D. Loughrey#, K. E. Watters#, A. Settle, J. B. LucksNucleic Acids Research. (2014). # = Equal contribution
Links: JournalPDF

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