Developing Novel L-RNA Aptamer to Inhibit Telomerase Activity and Regulate G-Quadruplex-Mediated Gene Expression

Description

RNA G-quadruplexes (rG4s) are non-classical RNA secondary structures that have been first observed decades ago. Over the years, these four-stranded structural motifs have been demonstrated to have significant regulatory roles in diverse biological processes, but challenges remain in targeting rG4 structures and understanding their biological consequences. To address this, the development of new rG4 targeting strategy and tool — especially the ability to target specific rG4 of interest, as well as the potential to interfere rG4-protein interaction and rG4-mediated biological events — are two crucial aspects for further exploring rG4 biology and developing novel approaches for rG4 targeting and gene regulation purposes. Our group have a strong and long-standing track record of detecting and targeting of rG4 structures. Recently, we have established transcript-specific and transcriptome-wide methods to report the formation and role of rG4s in the different biological systems, such as in human cells, plant and bacteria. Moreover, as proof-of-concept studies, we have developed the first series of rG4 targeting L-RNA aptamers and applied them to target rG4 of interest, of which the reported L-RNA aptamers demonstrated promising binding and selectivity properties. In this project, we will refine the rG4-SELEX strategy to establish a high-throughput platform, rG4-SELEX-seq to develop L-RNA aptamers to target rG4, convert the linear L-RNA aptamer into cyclic form via click chemistry to improve its rG4 binding and selectivity properties, and investigate the effect of cyclized L-RNA aptamer in controlling rG4-protein interaction and rG4-mediated gene expression. Our group have all the necessary equipment and expertise to execute the tasks in this proposal, and the new method and aptamer developed in this project, such as rG4-SELEX-seq, LRNA aptamer cyclization, and the L-RNA aptamer sequence itself will be patent-worthy. Specific rG4 targets of interest include the rG4 at the 5’ end of human telomerase RNA that is linked to telomere maintenance and genome instability, as well as the rG4 at the 3’UTR of amyloid precursor protein transcript, which is related to Alzheimer’s disease. Other biological important rG4s will also be studied. Our preliminary data are encouraging, and our group will continue to work closely together to ensure successful execution of this timely and exciting project. We anticipate the outcomes of this project will offer new insights into the understanding and controlling rG4 functions in biology.