There are as many as 100,000 different RNA molecules in the cell, most of which have unknown structure and function. A growing number of these have been directly connected with human disease. These RNAs interact with each other and with proteins, and control gene expression and cellular pathways in complex ways. Assessing amounts, locations, sequences and structures of these RNAs will become increasingly important to medicine as well as basic biology. Inspired by these facts, we are developing small-molecule tools to help researchers study the functions of RNAs.<br/><br/>As a strategy for mapping RNA structures in vivo, we have developed cell-permeable reagents that contain reactive acylimidazole groups. These enter the cell and acylate 2’-hydroxyl groups at positions where they are accessible to solvent. The acylation allows mapping of structure by RT-mediated primer extension. Combining this with RNA Seq methodologies has allowed us with collaborators to map the structures of large numbers of RNAs in mammalian cells, and this work is uncovering interesting data about protein-RNA interactions that occur in living systems.<br/><br/>As part of a strategy for bioorthogonal bond formation with RNAs and proteins, we are developing new water-soluble reagents and organocatalysts that enable rapid hydrazone and oxime formation. We have identified superior catalysts that foster bond formation efficiently at pH 7.4 and that have low toxicity to mammalian cells. We are using these compounds to develop novel strategies for labeling biomolecules in vivo. Notably, some of our catalysts can speed the reversal of bond formation with nucleic acids, and this is expected to lead to useful ways of unlocking biomedically important information from tissue samples worldwide.

Small-molecule strategies for mapping RNA structure and sequence

251st National Meeting (2016)

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