Poster abstracts

Poster number 121 submitted by Kaizheng Liu

Reversible Acylating Reagents to Dictate RNA’s Fate

Kaizheng Liu (Department of Chemistry, Carnegie Mellon University), Marta Rachwalak (Department of Chemistry, Carnegie Mellon University), Anna M. Kietrys (Department of Chemistry, Carnegie Mellon University)

RNA transcripts appear in various forms and play diverse roles in cellular processes[1,2]. New methods to manipulate RNA function and interactions increase the potential of revealing its new biochemical significances[3–5]. Recently, the 2′-OH acylation has gained wide attention due to its one-step procedure, high efficiency, and reversibility. Taking advantage of these findings, we have been working on the development of universal, effective, and biocompatible acylating reagents for understanding RNA interactome and gaining control over the biological function of various RNAs.
Herein, we present two approaches of molecular tools designed to acylate 2′-OH groups in single-stranded regions of RNA. The first strategy is based on the possibility of linking two RNA strains which would significantly disrupt RNA structure and function. Each probe contains two photocleavable groups connected by linkers of different chemical structures. Thanks to photosensitive units, the RNA function may be fully restored by UV light. The RNA will first be silenced by acylation in vitro, followed by its transfection into cells. To further develop this idea and find a fully biocompatible approach for in vivo applications, we have developed another set of reagents sensitive to enzyme. In these molecules, the disulfide bond acts as a thioredoxin reductase (TrxR) cleavable site, and self-immolative group enables the recovery of 2′-OH. Such design allows us to correlate the RNA activity with TrxR concentration. In cells with an elevated level of TrxR, treated RNA can be uncaged and alter gene expression. We have tested both approaches and optimized conditions for in vitro acylation of oligo RNAs. To show the potential biological application of proposed strategies, we plan to test these probes in cellulo. We believe that these molecules will serve as chemical tools to dictate RNA’s fate in cells. Additionally, designed reagents may help to map short and long-distance interactions in RNA complexes.

(1) Hsiao, K. Y.; Sun, H. S.; Tsai, S. J. Circular RNA – New Member of Noncoding RNA with Novel Functions. Exp Biol Med 2017, 242 (11), 1136–1141.
(2) Cech, T. R.; Steitz, J. A. The Noncoding RNA Revolution—Trashing Old Rules to Forge New Ones. Cell 2014, 157 (1), 77–94.
(3) Velema, W. A.; Park, H. S.; Kadina, A.; Orbai, L.; Kool, E. T. Trapping Transient RNA Complexes by Chemically Reversible Acylation. Angew Chem Int Ed Engl 2020, 59 (49), 22017–22022.
(4) Knutson, S. D.; Sanford, A. A.; Swenson, C. S.; Korn, M. M.; Manuel, B. A.; Heemstra, J. M. Thermoreversible Control of Nucleic Acid Structure and Function with Glyoxal Caging. J Am Chem Soc 2020, 142 (41), 17766–17781.
(5) Kadina, A.; Kietrys, A. M.; Kool, E. T.; Kadina, [ A; Ietrys, A. M. K.; Kool, E. T. RNA Cloaking by Reversible Acylation. Angewandte Chemie International Edition 2018, 57 (12), 3059–3063.

Keywords: RNA acylation, RNA Interactome , Gene regulation