Amine-substituted Indole Vinylquinoline As Fluorescent Probe For Specific Nucleic Acids

Nucleic acid with special structure is an essential polymeric biomolecule that carries various important biological functions in living cells such as coding,decoding,and controlling the expression of genes.Nucleic acid is mainly distributed in the nucleolus and cytoplasm of living cells because the nucleolus is the key site in the nucleus to synthesize and assemble ribosomal RNA(rRNA).The investigation of the morphology and functions of the nucleolus using RNA-specific fluorescent imaging technique in vivo are able to provide a powerful tool for cutting-edge researches in molecular biology,pathophysiology,and medical diagnostics.E36 is also an example of a dye that has the favorable physical characteristics of a RNA fluorescent probe.The structure is formed by conjugating the indolyl moiety with a 1-methylquinolinium scaffold though a ethylene bridge.The fluorescent ligand has good cell membrane permeability,counterstaining compatibility,and shows obvious selectivity to RNA.However,it still has some disadvantages of general water solubility and weak light stability.We hope to design RNA fluorescence probes with superior performance by changing the linking sites of the indolyl moiety on the basis of the E36 structure and introducing the amine side chain on the quinoline structure.The main work of the paper is:(1)The research progress of RNA and G-quadruplex fluorescent probes was reviewed,and the derivatives of amine-substituted indolyl vinyl quinolines were designed and synthesized,the compounds is formed by connecting the 2-position of quinoline and the 2-or 3-position of indol;(2)the fluorescence properties of the derivatives was studied by fluorescence spectroscopy.Compounds 1d and 2d were screened to specifically bind to RNA and can be used as a RNA fluorescent probes;(3)gel electrophoresis?cytotoxicity and cell imaging methods confirmed as RNA probes;(4)The fluorescence properties and detection applications of some derivatives as G-quadruplex probes were studied by UV spectroscopy,fluorescence spectroscopy,and cell imaging.The results indicate that the 3-ethylene bond indole selective effect on RNA specificly,especially compound 1d.However,compounds formed at the 2-position with vinyl groups show G-quadruplex selectivity.The amino group of conjugates could be significantly increased the water solubility of the compounds,due to the steric hindrance effect.the present study comprehensively demonstrated and compared the critical influence of a molecule structure one the RNA binding specificity against other types of nucleic acids with the small molecular ligands(1d,2d)that are built on a representative styryl ligand E36.The newly developed ligand with the structural consideration to enhance binding affinity and RNA selective exhibited much better performance than E36 in both in vitro and in vivo live cell imaging studies.In addition,the ligand is nuclear membrane permeable,particularly 1d,and shows excellent photostability and counterstaining property for real-time imaging or mapping of RNA and DNA distribution simultaneously in live cell experiments.The results obtained from this investigation are able provide useful indications for designing potent small molecular binders targeting RNA for the treatment of disease.