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Open Access Research

Global mRNA decay analysis at single nucleotide resolution reveals segmental and positional degradation patterns in a Gram-positive bacterium

Simen M Kristoffersen15, Chad Haase2, M Ryan Weil2, Karla D Passalacqua3, Faheem Niazi4, Stephen K Hutchison4, Brian Desany4, Anne-Brit Kolstø1, Nicolas J Tourasse1, Timothy D Read23* and Ole Andreas Økstad1*

Author Affiliations

1 Laboratory for Microbial Dynamics, Department of Pharmaceutical Biosciences, School of Pharmacy, University of Oslo, PB 1068 Blindern, 0316 Oslo, Norway

2 Department of Human Genetics, Division of Infectious Diseases, Emory University School of Medicine, 615 Michael Street, Atlanta, GA 30322-1013, USA

3 Department of Medicine, Division of Infectious Diseases, Emory University School of Medicine, 615 Michael Street, Atlanta, GA 30322-1013, USA

4 454 Life Sciences, A Roche Company, 1 Commercial Street, Branford, CT 06405, USA

5 PHARMAQ AS, PO Box 267 Skøyen, N-0213 Oslo, Norway

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Genome Biology 2012, 13:R30  doi:10.1186/gb-2012-13-4-r30

Published: 26 April 2012

Abstract

Background

Recent years have shown a marked increase in the use of next-generation sequencing technologies for quantification of gene expression (RNA sequencing, RNA-Seq). The expression level of a gene is a function of both its rate of transcription and RNA decay, and the influence of mRNA decay rates on gene expression in genome-wide studies of Gram-positive bacteria is under-investigated.

Results

In this work, we employed RNA-Seq in a genome-wide determination of mRNA half-lives in the Gram-positive bacterium Bacillus cereus. By utilizing a newly developed normalization protocol, RNA-Seq was used successfully to determine global mRNA decay rates at the single nucleotide level. The analysis revealed positional degradation patterns, with mRNAs being degraded from both ends of the molecule, indicating that both 5' to 3' and 3' to 5' directions of RNA decay are present in B. cereus. Other operons showed segmental degradation patterns where specific ORFs within polycistrons were degraded at variable rates, underlining the importance of RNA processing in gene regulation. We determined the half-lives for more than 2,700 ORFs in B. cereus ATCC 10987, ranging from less than one minute to more than fifteen minutes, and showed that mRNA decay rate correlates globally with mRNA expression level, GC content, and functional class of the ORF.

Conclusions

To our knowledge, this study presents the first global analysis of mRNA decay in a bacterium at single nucleotide resolution. We provide a proof of principle for using RNA-Seq in bacterial mRNA decay analysis, revealing RNA processing patterns at the single nucleotide level.