mRNA cap analogues substituted in the tetraphosphate chain with CX2: identification of O-to-CCl2 as the first bridging modification that confers resistance to decapping without impairing translation.

Autor:	Rydzik AM; Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Zwirki i Wigury 93, 02-089 Warsaw, Poland.; Chemistry Research Laboratory, University of Oxford, 12 Mansfield Road, Oxford OX1 3TA, UK., Warminski M; Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Zwirki i Wigury 93, 02-089 Warsaw, Poland., Sikorski PJ; Centre of New Technologies, University of Warsaw, Banacha 2c, 02-097 Warsaw, Poland., Baranowski MR; Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Zwirki i Wigury 93, 02-089 Warsaw, Poland., Walczak S; Centre of New Technologies, University of Warsaw, Banacha 2c, 02-097 Warsaw, Poland.; College of Inter-Faculty Individual Studies in Mathematics and Natural Sciences, University of Warsaw, Zwirki i Wigury 93, 02-089 Warsaw, Poland., Kowalska J; Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Zwirki i Wigury 93, 02-089 Warsaw, Poland., Zuberek J; Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Zwirki i Wigury 93, 02-089 Warsaw, Poland., Lukaszewicz M; Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Zwirki i Wigury 93, 02-089 Warsaw, Poland., Nowak E; Laboratory of Protein Structure, International Institute of Molecular and Cell Biology, 4 Ksiecia Trojdena Street, 02-109 Warsaw, Poland., W Claridge TD; Chemistry Research Laboratory, University of Oxford, 12 Mansfield Road, Oxford OX1 3TA, UK., Darzynkiewicz E; Division of Biophysics, Institute of Experimental Physics, Faculty of Physics, University of Warsaw, Zwirki i Wigury 93, 02-089 Warsaw, Poland.; Centre of New Technologies, University of Warsaw, Banacha 2c, 02-097 Warsaw, Poland., Nowotny M; Laboratory of Protein Structure, International Institute of Molecular and Cell Biology, 4 Ksiecia Trojdena Street, 02-109 Warsaw, Poland., Jemielity J; Centre of New Technologies, University of Warsaw, Banacha 2c, 02-097 Warsaw, Poland.
Jazyk:	angličtina
Zdroj:	Nucleic acids research [Nucleic Acids Res] 2017 Sep 06; Vol. 45 (15), pp. 8661-8675.
DOI:	10.1093/nar/gkx569
Abstrakt:	Analogues of the mRNA 5'-cap are useful tools for studying mRNA translation and degradation, with emerging potential applications in novel therapeutic interventions including gene therapy. We report the synthesis of novel mono- and dinucleotide cap analogues containing dihalogenmethylenebisphosphonate moiety (i.e. one of the bridging O atom substituted with CCl2 or CF2) and their properties in the context of cellular translational and decapping machineries, compared to phosphate-unmodified and previously reported CH2-substituted caps. The analogues were bound tightly to eukaryotic translation initiation factor 4E (eIF4E), with CCl2-substituted analogues having the highest affinity. When incorporated into mRNA, the CCl2-substituted dinucleotide most efficiently promoted cap-dependent translation. Moreover, the CCl2-analogues were potent inhibitors of translation in rabbit reticulocyte lysate. The crystal structure of eIF4E in complex with the CCl2-analogue revealed a significantly different ligand conformation compared to that of the unmodified cap analogue, which likely contributes to the improved binding. Both CCl2- and CF2- analogues showed lower susceptibility to hydrolysis by the decapping scavenger enzyme (DcpS) and, when incorporated into RNA, conferred stability against major cellular decapping enzyme (Dcp2) to transcripts. Furthermore, the use of difluoromethylene cap analogues was exemplified by the development of 19F NMR assays for DcpS activity and eIF4E binding. (© The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.)
Databáze:	MEDLINE
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