Identification of bacterial sRNA regulatory targets using ribosome profiling.

Autor:	Wang J; Wadsworth Center, New York State Department of Health, Albany, NY 12208, USA., Rennie W; Wadsworth Center, New York State Department of Health, Albany, NY 12208, USA., Liu C; Wadsworth Center, New York State Department of Health, Albany, NY 12208, USA., Carmack CS; Wadsworth Center, New York State Department of Health, Albany, NY 12208, USA., Prévost K; RNA Group, Department of Biochemistry, University of Sherbrooke, Sherbrooke, Quebec, J1H 5N4, Canada., Caron MP; RNA Group, Department of Biochemistry, University of Sherbrooke, Sherbrooke, Quebec, J1H 5N4, Canada., Massé E; RNA Group, Department of Biochemistry, University of Sherbrooke, Sherbrooke, Quebec, J1H 5N4, Canada., Ding Y; Wadsworth Center, New York State Department of Health, Albany, NY 12208, USA Department of Biomedical Sciences, University at Albany, Albany, NY 12201, USA., Wade JT; Wadsworth Center, New York State Department of Health, Albany, NY 12208, USA Department of Biomedical Sciences, University at Albany, Albany, NY 12201, USA joseph.wade@health.ny.gov.
Jazyk:	angličtina
Zdroj:	Nucleic acids research [Nucleic Acids Res] 2015 Dec 02; Vol. 43 (21), pp. 10308-20. Date of Electronic Publication: 2015 Nov 05.
DOI:	10.1093/nar/gkv1158
Abstrakt:	Bacteria express large numbers of non-coding, regulatory RNAs known as 'small RNAs' (sRNAs). sRNAs typically regulate expression of multiple target messenger RNAs (mRNAs) through base-pairing interactions. sRNA:mRNA base-pairing often results in altered mRNA stability and/or altered translation initiation. Computational identification of sRNA targets is challenging due to the requirement for only short regions of base-pairing that can accommodate mismatches. Experimental approaches have been applied to identify sRNA targets on a genomic scale, but these focus only on those targets regulated at the level of mRNA stability. Here, we utilize ribosome profiling (Ribo-seq) to experimentally identify regulatory targets of the Escherichia coli sRNA RyhB. We not only validate a majority of known RyhB targets using the Ribo-seq approach, but also discover many novel ones. We further confirm regulation of a selection of known and novel targets using targeted reporter assays. By mutating nucleotides in the mRNA of a newly discovered target, we demonstrate direct regulation of this target by RyhB. Moreover, we show that Ribo-seq distinguishes between mRNAs regulated at the level of RNA stability and those regulated at the level of translation. Thus, Ribo-seq represents a powerful approach for genome-scale identification of sRNA targets. (© The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.)
Databáze:	MEDLINE
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