Abstract :
[en] Non-canonical nucleic acid structures, such as G-quadruplexes, have recently attracted significant research efforts for their potential roles in the regulation of numerous biological processes [1]. There is now evidence that these specific nucleic acid structures modulate gene expression. The formation of hybrid DNA:RNA G-quadruplexes (HQs) where each strand contribute to the structure with two series of guanines, was recently found to form in DNA during transcription [2]. It has been evidenced that the HQ formation competes with intramolecular G-quadruplexe motifs (DQ) and plays a major role in regulating transcription [3]. Given their very recent discovery, only a few studies on the formation of these hybrid structures have been conducted so far. However, the knowledge of their mechanism of formation and their detailed structure is important for understanding their biological function, as well as for manipulating gene expression by targeting HQs.
We propose here to study these hybrid DNA:RNA G-quadruplexe structures by AFM-based single molecule force spectroscopy to obtain detailed information on their formation mechanism and dynamics, at the single molecule level. We investigated the structure and the interactions that govern these complexes, their mechanical stability and their lability. We also studied the DNA:DNA G-quadruplexes to compare the forces and stretching profile.
In order to study the bimolecular G-quadruplexes by AFM-based force spectroscopy, one partner must be grafted on the probe and the other on the surface. By performing approach-retraction cycles, we were able to identify the specific unfolding profile of these structures, which is a saw-tooth pattern, showing the consecutive breakings of intramolecular interactions. We have highlighted the most probable force values necessary to break these interactions and the associated lengths revealed consecutively to these breakings.
Our study should contribute to understand the effect of the spatial proximity between DNA and RNA strands on the structures that can be formed, on their kinetics of formation, and on their resistance to mechanical forces.
References
[1] M. L. Bochman, K. Paeschke, V. A. Zakian, Nat. Rev. Genet., 2012, 13, 770-780.
[2] K. W. Zheng, S. Xiao, J. Q. Liu, J. Y. Zhang, Y. H. Hao, Z. Tan, Nucleic Acids Res., 2013, 41, 5533-5541.
[3] R.-Y. Wu, K.-W Zheng, J.-Y. Zhang, Y.-H. Hao, Z. Tan, Angew. Chem. Int. Ed., 2015, 54, 2477-2481.
