DNA Structural Polymorphism And Its Recognition And Regulation By Small Molecules

Nucleic acids, key genetic materials in biological systems, can form a very widerange of different structures, aside from the well-known DNA double helix. Thisdouble helix is highly unusual in which the structure is largely independent of thesequence. For other structures, what form they adopt, and how stably, is controlled bytheir sequences as well as in particular by the different chemical properties of thenucleobases. The intriguing structural diversity in folded topologies available toguanine-rich nucleic acid repeat sequences have made four-stranded G-quadruplexstructures the focus of both basic and applied research as growing evidence suggeststhat these structures may play important biological roles in vivo. Distributed widely inthe human genome as targets for regulating gene expression and chromosomalmaintenance, they offer unique avenues for future cancer drug development. Forinstance, human telomeric DNAs are non-coding highly repetitive sequencesconsisting of tandem repeats of the sequence d(TTAGGG) that are relevant to cellaging and cancer. Thus, G-quadruplexes formed by human telomeric DNA sequenceshave been the subject of increasing attention due to their potential as a therapeutictarget for cancer. G-quadruplexes which are also prevalent in non-telomeric portionsof the genome, particularly in gene promoters, where quadruplex formation must be inaccordance with duplex dissociation, can also regulate the transcription of someoncogenes, suggesting an additional anticancer potential. However, althoughG-quadruplex nucleic acids are considered as greatly significant roles in biologicalprocesses, the existence of these structures in living cells still lack efficiently directevidence and what conformation they adopt under cellular conditions is totally unclear.Thus, elucidating their structures in cellular conditions, and further detecting andregulating their structural behaviors by manual pathways are critical issues at presentand undoubtedly deserve wide investigations.One research field of my thesis is to elucidate DNA structural polymorphismunder different environments in order to reveal their real behaviors in living cells. DNA structures could be influenced by molecular crowding and dehydration that bothlikely occur in living cells. Previous analyses usually considered the static state or thestable state of DNA structures, while we provided a new insight into the dynamicconversion of the human telomere sequence under molecular crowding conditions atphysiologic temperature. As the cellular environment is crowded by macromolecules,the results here may correspond with the behavior of human telomere DNA in livingcells. Besides, I also explored DNA structures in hydrophobic circumstances andobserved that unwinding activities of DNA duplexes by helicase would be stronglyinfluenced by dehydration effect, providing a novel insight for duplex dissociationand G-quadruplex formation during gene transcription as enzyme could createhydrophobic pockets in which the duplexes would be located.The other focus of my thesis is to investigate interactions between DNA andsynthesized molecules in order to develop new anticancer drugs and bio-detectingagents. We designed and generated a dinuclear ruthenium complex that couldspecifically and selectively target DNA G-quadruplex. Fluorescent experimentsdemonstrated that this novel dinuclear ruthenium (II) complex had high fluorescenceresponse when interacting with G-quadruplex structures, with high selectivity overduplex DNA at the same time. Direct observation of G-quadruplex structures bynaked eyes both in vitro and in vivo was also realized. Detecting G-quadruplexstructures has great significance for cell proliferation, cancer research, and drugdevelopment, and this molecule is a quite promising tool. We also report a novelsimple small molecule (pyridyl carboxamide), consisting of three pyridine rings andfour amide bonds. Comparing with some reported G-quadruplex ligands, thismolecule not only induced a slow kinetic unimolecular human telomericG-quadruplex transformation process, further revealing the structural polymorphismof human telomeric DNA, but also regulate duplex-quadruplex competition even ifthe G-rich region was trapped in long double-stranded DNA. It is widely believed thatthe dissociation of duplexes is involved in gene transcription and that the formation ofthe G-quadruplex influences some oncogene expression. Py-Am exhibited strongG-quadruplex-forming ability within a long dsDNA sequence, suggesting it would have potent effects on the G-quadruplex-forming sequences involved in genetranscription.Overall, this work we studied here on DNA structural polymorphism forrevealing their real behaviors in living cells as well as on construction of new smallmolecules that can target G-quadruplexes for developing new bio-detecting agentsand anticancer drugs will provide new considerations and approaches to explorepractical applications of manual tools.