Uncovering How RNA Molecules ‘Make Decisions’ On the Fly: Towards Understanding and Engineering CoTranscriptional RNA Folding

RNAs are emerging as a powerful substrate for engineering gene expression and cellular behavior since they are now known to control almost all aspects of gene expression. As with all biomolecules, RNA function is intimately related to its structure, since RNA can adopt structures that selectively modulate gene expression. Central questions in biology and bioengineering then are: How do RNAs fold inside cells?; and How can we engineer these folds to control gene expression? In this talk, I will present our work at the interface of these two questions and share results that are beginning to uncover design principles for engineering RNAs for an array of applications, ranging from metabolic engineering to diagnostics.

I will start by presenting our work on engineering RNA molecular switches that control transcription. The desire to uncover design principles for engineering these RNAs motivates our development of SHAPE-Seq, a technology that couples chemical probing with next-generation sequencing and that helps characterize RNA structures on an ‘omics’ scale. I will then describe our exciting recent developments in using SHAPE-Seq to help break open one of the final frontiers of RNA structure-function relationships by uncovering at nucleotide resolution how RNAs fold cotranscriptionally. Specifically I will highlight new data on uncovering the ligand-dependent folding pathways of riboswitches. This new ability is allowing us to ask deep questions about how RNA molecules make regulatory decisions ‘on the fly’ during the dynamic process of transcription. By probing the fundamental processes of RNA folding and function, these studies are expected to greatly aid RNA engineering.