A ratiometric fluorescence RRE RNA-targeted assay for a new fluorescence ligand

• Published in: Biosensors & bioelectronics 2016-12, Vol.86, p.287-292
• Main Authors: Qi, Liang, Wei, Jing-Ru, Lv, Xiao-Jun, Huo, Yuan, Zhang, Zhi-Qi
• Format: Article
• Language: English
• Subjects: Assaying; Cadmium tellurides; Construction; Equipment Design; Equipment Failure Analysis; Fluorescence; Human immunodeficiency virus 1; ICR 191; Lentivirus; Ligand-RNA interaction; Ligands; Peptides; Quantum Dots; Ratiometric fluorescence assay; Reproducibility of Results; Response Elements - genetics; Rev Protein; Ribonucleic acids; RNA - chemistry; RNA - genetics; RRE RNA; Sensitivity and Specificity; Spectrometry, Fluorescence - methods
• ISSN: 0956-5663; 1873-4235
• DOI: 10.1016/j.bios.2016.06.051

The Rev protein regulates HIV-1 gene expression. Small ligands that bind to the Rev response element (RRE) RNA would inhibit Rev function and suppress HIV-1 replication. A novel ratiometric fluorescence assay was applied in the present study to monitor ligand-RNA interactions by using red-emitting CdTe quantum dots (QDs) coated with silica as a reference. A small fluorescence ligand, ICR 191, was found to interact with RRE at the Rev binding site and compete with the Rev peptide. After adding red-emitting QDs to the interaction system, it was observed that ICR 191 did not fluoresce upon the addition of RRE, and fluorescence recovered when ICR 191 was displaced by a Rev model peptide, whereas the fluorescence of QDs remained constant. Furthermore, variations in the fluorescence ratios between ICR 191 and QDs were exploited to characterize the interactions of Rev with two known antagonists, neomycin B and tobramycin, by using RRE RNA with ICR 191 as a fluorescence indicator. Together, our results demonstrated that ratiometric fluorescence-based nanotechnology applications can be used for ligand−RNA interaction assays. This ICR 191-RRE RNA interaction assay can potentially be developed to build a screening model for assessing antagonists of the Rev binding element in RRE. •A new small fluorescence ligand was found to interact with RRE at the Rev binding site.•The ratiometric fluorescence assay was first applied to monitor ligand-RNA interactions.•ICR 191 as a fluorescent indicator is potentially used to screen Rev antagonists.