RRM
2013 Rustbelt RNA Meeting
RRM
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Poster abstracts
Abstract:
A library screening approach identified naturally occurring sequences for an RNA G-quadruplex binding ligand
Abstract
G-quadruplex (GQ) structures are secondary structures formed by both RNA and DNA. It consists of four guanines joined by Hoogsteen base pairings and stabilized by a monovalent cation.1 RNA G-quadruplexes have gained an increasing attention due to their critical roles in translational regulation of many important genes such as VEGEF, NRAS, FMRI and etc.2,3,4 Although designing of small molecules to target RNA GQs have become an important therapeutic approach, it is difficult to achieve the selectivity among GQs formed by the various RNA sequences. Therefore, it is crucial to identify all possible GQs in the human transcriptome that can potentially bind to these small molecules. A pilot study was performed to address this issue. We designed an RNA sequence library which has the possibility to form thousands of different GQ structures. Then we screened this library against kanamycin which is known to bind with RNA secondary structures including GQs.5 We found seven different RNA sequences that bind selectively with kanamycin and three of them are found in human transcriptome. The formation of GQ by each RNA sequence was confirmed by circular dichroism (CD), RNase T1 footprinting and CD melting. The binding affinities of kanamycin with the selected RNAs were compared using fluorescence Intercalator displacement assay (FID) where a fluorescence dye is replaced from the GQ by titrating with kanamycin.6 These studies show the possible GQ targets for kanamycin and how tightly such sequences interact with it. Therefore our studies showed the importance of this library screening approach to identify possible G-quadruplex targets for a small molecule which can be used as therapeutic. Elaborating the pool of the G-quadruplex library will be a further improvement of this approach to identify all potential targets in human transcriptome for a given small drug molecule.
References:
References
1. S. Neidle and Balasubramanian, S. , RSC Biomolecular Sciences., 2006.
2. J. C. Darnell, K. B. Jensen, P. Jin, V. Brown, S. T. Warren and R. B. Darnell, Cell, 2001, 107, 489-499.
3. M. J. Morris, Y. Negishi, C. Pazsint, J. D. Schonhoft and S. Basu, J. Am. Chem. Soc., 2010, 132, 17831-17839.
4. S. Kumari, A. Bugaut, J. L. Huppert and S. Balasubramanian, Nat. Chem. Biol., 2007, 3, 218-221.
5. O. Aminova, D. J. Paul, J. L. Childs-Disney and M. D. Disney, Biochemistry, 2008, 47, 12670-12679.
6. D. Monchaud, C. Allain and M. P. Teulade-Fichou, Bioorg. Med. Chem. Lett., 2006, 16, 4842-4845.
Keywords: RNA G-quadruplex, G-quadruplex Ligands , Library Screening
