Design and Synthesis of Caged Fluorescent Nucleotides and Application to Live-cell RNA Imaging

• Published in: Chembiochem : a European journal of chemical biology Vol. 12; no. 18; pp. 2871 - 2880
• Main Authors: Ikeda, Shuji, Kubota, Takeshi, Wang, Dan Ohtan, Yanagisawa, Hiroyuki, Umemoto, Tadashi, Okamoto, Akimitsu
• Format: Journal Article
• Language: English
• Published: Weinheim WILEY-VCH Verlag 16.12.2011; WILEY‐VCH Verlag
• Subjects: Drug Design; Fluorescence; fluorescence spectroscopy; fluorescent probes; nucleotides; Nucleotides - chemistry; photochemistry; photolysis; RNA - chemistry; RNA recognition
• ISSN: 1439-4227; 1439-7633
• DOI: 10.1002/cbic.201100523

A binary photocontrolled nucleic acid probe that contains a nucleotide modified with one photolabile nitrobenzyl unit and two hybridization‐sensitive thiazole orange units has been designed for area‐specific fluorescence imaging of RNA in a cell. The synthesized probe emitted very weak fluorescence regardless of the presence of the complementary RNA, whereas it showed hybridization‐sensitive fluorescence emission at 532 nm after photoirradiation at 360 or 405 nm for uncaging. Fluorescence suppression of the caged probe was attributed to a decrease in the duplex‐formation ability. Caged fluorescent nucleotides with other emission wavelengths (622 and 724 nm) were also synthesized in this study; they were uncaged by 360 nm irradiation, and emitted fluorescence in the presence of the complementary RNA. Such probes were applied to area‐specific RNA imaging in a cell. Only probes in the defined irradiation area were activated by uncaging irradiation, and subnuclear mRNA diffusion in a living cell was monitored. A binary photocontrolled nucleic acid probe that contained a nucleotide modified with one photolabile unit and two hybridization‐sensitive fluorescent dyes has been designed for area‐specific fluorescence imaging of RNA in a cell. Only probes in the defined irradiation area were activated by uncaging irradiation; subnuclear mRNA diffusion in a living cell was monitored.