title: Synthetic Capped mRNAs for Cap-Specific Photo-Cross-Linking Experiments.
abstract: The 7-methylguanosine triphosphate cap present at the 5' ends of eukaryotic mRNAs plays numerous roles in mRNA expression and metabolism. The identification and studies on cap-binding partners can be significantly advanced using tailored chemical tools such as synthetic cap analogues or RNAs carrying modified cap structures. Here we provide protocols for the production of mRNAs specifically labeled within the 5' cap with a nucleoside capable of being photo-activated, either 6-thioguanosine or 7-methyl-6-thioguanosine, which can be used in photo-cross-linking experiments to identify or characterize cap-binding biomolecules. We also describe a protocol for the cross-linking experiments with capped RNAs to map histone H4 cap-binding pocket.
authors.affiliation: Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Zwirki i Wigury 93, Warsaw, 02-089, Poland.
authors.forename: Joanna
authors.initials: J
authors.lastname: Kowalska
authors.affiliation: Architecture et Réactivité de l'ARN, Université de Strasbourg, CNRS, Institut de Biologie Moléculaire et Cellulaire, 15 rue René Descartes, 67084, Strasbourg CEDEX, France. f.martin@ibmc-cnrs.unistra.fr.
authors.forename: Franck
authors.initials: F
authors.lastname: Martin
authors.affiliation: Center of New Technologies, University of Warsaw, Banacha 2c, 02-097, Warsaw, Poland. jacekj@biogeo.uw.edu.pl.
authors.forename: Jacek
authors.initials: J
authors.lastname: Jemielity
mesh_terms: D000818:Animals; D003432:Cross-Linking Reagents; D006151:Guanosine / Q000031:analogs & derivatives* / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D006801:Humans; D015394:Molecular Structure; D014176:Protein Biosynthesis; D012314:RNA Cap Analogs / Q000737:chemistry / Q000378:metabolism*; D012333:RNA, Messenger / Q000737:chemistry* / Q000378:metabolism; D013872:Thionucleosides / Q000378:metabolism*; D014466:Ultraviolet Rays
chemical_list: D003432:Cross-Linking Reagents; D006657:Histones; D012314:RNA Cap Analogs; D012333:RNA, Messenger; D013872:Thionucleosides; D006151:Guanosine; C017516:7-methylguanosine triphosphate; C005783:6-thioguanosine
keywords: 5′ cap analogs; 6-thioguanosine; Cap binding; Histone mRNA; Photo-cross-linking; Transcription; mRNA