Poster abstracts
Poster number 79 submitted by Marta Rachwalak
A key to understanding RNA’s interactome: photocleavable probes for spatiotemporal control of RNA
Marta Rachwalak (Chemistry, Carnegie Mellon University), Avleen K. Chawla (Chemistry, Carnegie Mellon University), Megan L. Van Horn (Chemistry, Carnegie Mellon University), Anna M. Kietrys (Chemistry, Carnegie Mellon University)
Abstract:
Caging is one of the key methods for imaging and studying interactions and structural aspects of RNA inside the cell. Despite the rapid improvements in caging and cloaking techniques, scientists still struggle to develop universal and effective tools for understanding the complexity and dynamics of the RNA interactome in cellulo. To overcome this absence of appropriate tools, we designed and synthesized a set of photoreversible probes for studying RNA’s function and interactions within cells. An introduction of photocleavable protecting groups (PPGs) ensures control over the biological function of various RNAs (e.g., siRNA, mRNA) and allows for structure and function reactivation, in a spatiotemporal manner, in the living cell. Using the same set of probes in living cells may help to map short- and long-distance interactions in RNA complexes.
Herein, we present a synthesis of photocleavable (INPhRNA) probes designed to block RNA’s biological function through acylation of 2′-OH groups in single stranded regions. Protection of 2’-OH moieties significantly disrupts RNA structure and as such its function. The RNA function may be fully restored via UV-irradiation without any cellular damage. The photosensitive residue of the probes consists of PPG units connected by linkers. By modulating the type of linker and its length, we are able to affect both the solubility and reactivity of the probe. To show the biological applicability of the proposed strategy, we tested the probe’s cytotoxicity and potential for spatiotemporal control over siRNA and mRNA. We believe that the developed caging probes may serve as an efficient method for postsynthetic modification of RNA to protect them from degradation and direct their cellular activity. Additionally, they may be used as tools to explore RNA interactome complexity in vivo.
Keywords: RNA, photocleavable protecting group, RNA interactome