This Program Project Grant brings together seven principal investigators committed to understanding the principles by which RNA molecules fold into their biologically active structures. The proposal comprises four Projects and a Resource Core, all bound together by a central hypothesis: RNA folding can be understood and eventually predicted from a reductionist perspective. RNA looked like a simple messenger in the early days of structural and molecular biology. The last two decades have demonstrated diverse biological, chemical, and physical roles of RNA, and yet our mechanistic understanding remains limited. Deep investigation of the physical behavior of RNA will enable a full description of gene expression and its control. In particular, a predictive and quantitative understanding will catalyz rational RNA engineering at the molecular and systems biology levels and ultimately facilitate rational therapeutic interventions that target or involve RNA. Our component projects build from simple to complex. Project 1 studies the ion atmosphere that surrounds simple helices and uses an integrated experimental-computational feedback loop to develop an understanding of electrostatics, the largest and most general force influencing the behavior and interactions of RNA. Project 2 focuses on helix-junction-helix motifs, the building blocks of RNA tertiary structure. We will develop and apply novel experimental and analytical approaches to fully describe the conformational ensemble for these elements. Project 3 determines the conformational ensembles of RNAs in their unfolded, folded, and intermediate states and defines the energetics of the transitions between these states. We will compare the behaviors of P4-P6 RNA and RNA/protein complexes to the behavior predicted from the properties of their elements, as delineated from Project 2. We thereby directly test the hypothesis that the energetics and behavior of complex RNAs can be understood from the 'sum of their parts', a hypothesis that if true, will lead to a general ability to predict RNA folding and energetics. Projct 4 builds on the reductionist framework developed in the other projects to now address RNA folding kinetics, testing quantitative predictions and developing novel systematic approaches to dissect the influence of co-transcription folding. Together, these studies will uncover basic properties and behaviors that define the physical, chemical, and biological behavior of RNA and catalyze a transformation from a descriptive to quantitative predictive understanding of RNA.
HEALTH RELEVANCE: RNA is the central component in gene expression, transmitting information from DNA's stored genetic code. But RNA molecules are also dynamic and structured entities, and interactions with them are critical in development and in the regulation o all life forms. Thus, aberrant behavior of RNA can cause disease, and, conversely, the control of RNA-mediated processes in pathogens provides potential routes to new therapies.
| Liu, Bei; Merriman, Dawn K; Choi, Seung H et al. (2018) A potentially abundant junctional RNA motif stabilized by m6A and Mg2. Nat Commun 9:2761 |
| Denny, Sarah Knight; Bisaria, Namita; Yesselman, Joseph David et al. (2018) High-Throughput Investigation of Diverse Junction Elements in RNA Tertiary Folding. Cell 174:377-390.e20 |
| Kimsey, Isaac J; Szymanski, Eric S; Zahurancik, Walter J et al. (2018) Dynamic basis for dG•dT misincorporation via tautomerization and ionization. Nature 554:195-201 |
| Merriman, Dawn K; Yuan, Jiayi; Shi, Honglue et al. (2018) Increasing the length of poly-pyrimidine bulges broadens RNA conformational ensembles with minimal impact on stacking energetics. RNA 24:1363-1376 |
| Gracia, Brant; Al-Hashimi, Hashim M; Bisaria, Namita et al. (2018) Hidden Structural Modules in a Cooperative RNA Folding Transition. Cell Rep 22:3240-3250 |
| Zettl, Thomas; Das, Rhiju; Harbury, Pehr A B et al. (2018) Recording and Analyzing Nucleic Acid Distance Distributions with X-Ray Scattering Interferometry (XSI). Curr Protoc Nucleic Acid Chem 73:e54 |
| Ganser, Laura R; Lee, Janghyun; Rangadurai, Atul et al. (2018) High-performance virtual screening by targeting a high-resolution RNA dynamic ensemble. Nat Struct Mol Biol 25:425-434 |
| Boyle, Evan A; Andreasson, Johan O L; Chircus, Lauren M et al. (2017) High-throughput biochemical profiling reveals sequence determinants of dCas9 off-target binding and unbinding. Proc Natl Acad Sci U S A 114:5461-5466 |
| Bisaria, Namita; Jarmoskaite, Inga; Herschlag, Daniel (2017) Lessons from Enzyme Kinetics Reveal Specificity Principles for RNA-Guided Nucleases in RNA Interference and CRISPR-Based Genome Editing. Cell Syst 4:21-29 |
| Gleitsman, Kristin R; Sengupta, Raghuvir N; Herschlag, Daniel (2017) Slow molecular recognition by RNA. RNA 23:1745-1753 |
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