title: RNA Hairpin Folding in the Crowded Cell.
abstract: Precise secondary and tertiary structure formation is critically important for the cellular functionality of ribonucleic acids (RNAs). RNA folding studies were mainly conducted in vitro, without the possibility of validating these experiments inside cells. Here, we directly resolve the folding stability of a hairpin-structured RNA inside live mammalian cells. We find that the stability inside the cell is comparable to that in dilute physiological buffer. On the contrary, the addition of in vitro artificial crowding agents, with the exception of high-molecular-weight PEG, leads to a destabilization of the hairpin structure through surface interactions and reduction in water activity. We further show that RNA stability is highly variable within cell populations as well as within subcellular regions of the cytosol and nucleus. We conclude that inside cells the RNA is subject to (localized) stabilizing and destabilizing effects that lead to an on average only marginal modulation compared to diluted buffer.
authors.affiliation: Physical Chemistry II, Ruhr-Universität Bochum, Universitätsstr. 150, 44801, Bochum, Germany.
authors.forename: Mimi
authors.initials: M
authors.lastname: Gao
authors.affiliation: Physical Chemistry II, Ruhr-Universität Bochum, Universitätsstr. 150, 44801, Bochum, Germany.
authors.forename: David
authors.identifier: http://orcid.org/0000-0001-9393-503X
authors.initials: D
authors.lastname: Gnutt
authors.affiliation: Institute for Biochemistry, Ernst-Moritz-Arndt-University Greifswald, Felix-Hausdorff-Str. 4, 17487, Greifswald, Germany.
authors.forename: Axel
authors.initials: A
authors.lastname: Orban
authors.affiliation: Institute for Biochemistry, Ernst-Moritz-Arndt-University Greifswald, Felix-Hausdorff-Str. 4, 17487, Greifswald, Germany.
authors.forename: Bettina
authors.initials: B
authors.lastname: Appel
authors.affiliation: Microbial Biology, Ruhr-Universität Bochum, Universitätsstr. 150, 44801, Bochum, Germany.
authors.forename: Francesco
authors.initials: F
authors.lastname: Righetti
authors.affiliation: Physical Chemistry I, Technical University Dortmund, Otto-Hahn-Str. 4a, 44227, Dortmund, Germany.
authors.forename: Roland
authors.initials: R
authors.lastname: Winter
authors.affiliation: Microbial Biology, Ruhr-Universität Bochum, Universitätsstr. 150, 44801, Bochum, Germany.
authors.forename: Franz
authors.initials: F
authors.lastname: Narberhaus
authors.affiliation: Institute for Biochemistry, Ernst-Moritz-Arndt-University Greifswald, Felix-Hausdorff-Str. 4, 17487, Greifswald, Germany.
authors.forename: Sabine
authors.initials: S
authors.lastname: Müller
authors.affiliation: Physical Chemistry II, Ruhr-Universität Bochum, Universitätsstr. 150, 44801, Bochum, Germany. simon.ebbinghaus@rub.de.
authors.forename: Simon
authors.identifier: http://orcid.org/0000-0001-9309-1279
authors.initials: S
authors.lastname: Ebbinghaus
mesh_terms: D031541:Fluorescence Resonance Energy Transfer; D009690:Nucleic Acid Conformation; D011092:Polyethylene Glycols / Q000737:chemistry; D058105:Polymerization; D012313:RNA / Q000737:chemistry*
chemical_list: D011092:Polyethylene Glycols; D012313:RNA
keywords: RNA; biophysics; folding stability; in-cell spectroscopy; macromolecular crowding
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