title: Chemical probing of RNA in living cells.
abstract: RNAs need to adopt a specific architecture to exert their task in cells. While significant progress has been made in describing RNA folding landscapes in vitro, understanding intracellular RNA structure formation is still in its infancy. This is in part due to the complex nature of the cellular environment but also to the limited availability of suitable methodologies. To assess the intracellular structure of large RNAs, we recently applied a chemical probing technique and a metal-induced cleavage assay in vivo. These methods are based on the fact that small molecules, like dimethyl sulfate (DMS), or metal ions, such as Pb(2+), penetrate and spread throughout the cell very fast. Hence, these chemicals are able to modify accessible RNA residues or to induce cleavage of the RNA strand in the vicinity of a metal ion in living cells. Mapping of these incidents allows inferring information on the intracellular conformation, metal ion binding sites or ligand-induced structural changes of the respective RNA molecule. Importantly, in vivo chemical probing can be easily adapted to study RNAs in different cell types.
authors.affiliation: Department of Biochemistry and Cell Biology, Max F. Perutz Laboratories, University of Vienna, Vienna, Austria.
authors.forename: Michael
authors.initials: M
authors.lastname: Wildauer
authors.forename: Georgeta
authors.initials: G
authors.lastname: Zemora
authors.forename: Andreas
authors.initials: A
authors.lastname: Liebeg
authors.forename: Verena
authors.initials: V
authors.lastname: Heisig
authors.forename: Christina
authors.initials: C
authors.lastname: Waldsich
mesh_terms: D058645:Denaturing Gradient Gel Electrophoresis / Q000379:methods; D057809:HEK293 Cells; D006801:Humans; D007854:Lead / Q000378:metabolism*; D015336:Molecular Probe Techniques; D015335:Molecular Probes / Q000378:metabolism*; D009690:Nucleic Acid Conformation; D012313:RNA / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D013463:Sulfuric Acid Esters / Q000378:metabolism*; D015003:Yeasts / Q000166:cytology / Q000378:metabolism
chemical_list: D015335:Molecular Probes; D013463:Sulfuric Acid Esters; D007854:Lead; D012313:RNA; C007482:dimethyl sulfate