Affiliation:
1. Laboratory of Molecular Cell Dynamics, Faculty of Advanced Life Science, Hokkaido University , Sapporo , Japan
2. Experimental Biomolecular Physics, Department of Applied Physics, Royal Institute of Technology (KTH) , Stockholm , Sweden
Abstract
AbstractGuanine (G)-rich nucleic acids are prone to assemble into four-stranded structures, so-called G-quadruplexes. Abnormal GGGGCC repeat elongations, and in particular their folding states, are associated with amyotrophic lateral sclerosis and frontotemporal dementia. Due to methodological constraints however, most studies of G quadruplex structures are restricted to in vitro conditions. Evidence of how GGGGCC repeats form into G-quadruplexes in vivo is sparse. We devised a readout strategy, exploiting the sensitivity of trans-cis isomerization of cyanine dyes to local viscosity and sterical constraints. Thereby, folding states of cyanine-labeled RNA, and in particular G-quadruplexes, can be identified in a sensitive manner. The isomerization kinetics, monitored via fluorescence blinking generated upon transitions between a fluorescent trans isomer and a non-fluorescent cis isomer, was first characterized for RNA with GGGGCC repeats in aqueous solution using fluorescence correlation spectroscopy and transient state (TRAST) monitoring. With TRAST, monitoring the isomerization kinetics from how the average fluorescence intensity varies with laser excitation modulation characteristics, we could then detect folding states of fluorescently tagged RNA introduced into live cells.
Funder
Swedish Research Council
Knut and Alice Wallenberg Foundation
Swedish Foundation for Strategic Research
JSPS
Fostering Joint International Research
JSPS Grant-in-Aid for Scientific Research
Japan Agency for Medical Research and Development
Canon Foundation Research
Hoansha Foundation in Japan
Hokkaido University Office for Developing Future Research Leaders
Publisher
Oxford University Press (OUP)
Reference63 articles.
1. Potassium physiology;Thier;Am. J. Med.,1986
2. RNA G-quadruplexes in biology: principles and molecular mechanisms;Fay;J. Mol. Biol.,2017
3. Detecting RNA G-quadruplexes (rG4s) in the transcriptome;Kwok;Cold Spring Harb. Perspect. Biol.,2018
4. RNA G-quadruplexes: emerging mechanisms in disease;Cammas;Nucleic Acids Res.,2017
5. A comparison of DNA and RNA quadruplex structures and stabilities;Joachimi;Bioorg. Med. Chem.,2009
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