Bacterial Antisense RNAs: How Many Are There, and What Are They Doing?

Thomason, Maureen K.; Storz, Gisela

doi:10.1146/annurev-genet-102209-163523

Cited by 324 publications

(341 citation statements)

References 125 publications

(165 reference statements)

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“…To exclude the possibility that the aTSS were experimental artifacts of nonspecific priming during cDNA synthesis (33), the dRNAseq results were compared with available tiling array data (13) and with a transcriptome array in which RNA was labeled directly to avoid such artifacts. This comparison verified the existence of a plethora of antisense transcripts in this organism and recapitulates recent findings of massive antisense transcription in other prokaryotes (34). The compact genome of the human pathogen Helicobacter pylori transcribes asRNAs for 46% of all annotated ORFs (8), and in the archaeon Sulfolobus solfataricus 6.1% of all genes were associated with asRNAs (30).…”

Section: Discussionsupporting

confidence: 87%

An experimentally anchored map of transcriptional start sites in the model cyanobacterium Synechocystis sp. PCC6803

Mitschke

Georg

Scholz

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

350

459

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There has been an increasing interest in cyanobacteria because these photosynthetic organisms convert solar energy into biomass and because of their potential for the production of biofuels. However, the exploitation of cyanobacteria for bioengineering requires knowledge of their transcriptional organization. Using differential RNA sequencing, we have established a genome-wide map of 3,527 transcriptional start sites (TSS) of the model organism Synechocystis sp. PCC6803. One-third of all TSS were located upstream of an annotated gene; another third were on the reverse complementary strand of 866 genes, suggesting massive antisense transcription. Orphan TSS located in intergenic regions led us to predict 314 noncoding RNAs (ncRNAs). Complementary microarray-based RNA profiling verified a high number of noncoding transcripts and identified strong ncRNA regulations. Thus, ∼64% of all TSS give rise to antisense or ncRNAs in a genome that is to 87% protein coding. Our data enhance the information on promoters by a factor of 40, suggest the existence of additional small peptide-encoding mRNAs, and provide corrected 5′ annotations for many genes of this cyanobacterium. The global TSS map will facilitate the use of Synechocystis sp. PCC6803 as a model organism for further research on photosynthesis and energy research.gene expression regulation | promoter prediction | RNA polymerase

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Section: Discussionsupporting

confidence: 87%

An experimentally anchored map of transcriptional start sites in the model cyanobacterium Synechocystis sp. PCC6803

Mitschke

Georg

Scholz

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

350

459

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“…Instead diverse classes of RNA transcripts are now known to fulfill important functions that go far beyond serving as simple messengers for protein synthesis or as house-keeping molecules. In bacteria diverse non-coding RNA elements have been identified, including anti-sense RNAs (Thomason and Storz, 2010), riboswitches (Winkler and Breaker, 2005) or small RNAs (sRNAs; see below). In higher eukaryotes, as demonstrated by the Encyclopedia of DNA elements (ENCODE) project, protein-coding genes constitute only a very minor fraction (~3%) of the genome and are outnumbered by non-coding genes (Dunham et al, 2012).…”

Section: The Transcriptome As a Snapshot Of An Infected Cell's Physiomentioning

confidence: 99%

Dual RNA-seq of pathogen and host

2012

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SummaryThe infection of a eukaryotic host cell by a bacterial pathogen is one of the most intimate examples of cross-kingdom interactions in biology. Infection processes are highly relevant from both a basic research as well as a clinical point of view. Sophisticated mechanisms have evolved in the pathogen to manipulate the host response and vice versa host cells have developed a wide range of anti-microbial defense strategies to combat bacterial invasion and clear infections.However, it is this diversity and complexity that makes infection research so challenging to technically address as common approaches have either been optimized for bacterial or eukaryotic organisms. Instead, methods are required that are able to deal with the often dramatic discrepancy between host and pathogen with respect to various cellular properties and processes. One class of cellular macromolecules that exemplify this host-pathogen heterogeneity is given by their transcriptomes: Bacterial transcripts differ from their eukaryotic counterparts in many aspects that involve both quantitative and qualitative traits. The entity of RNA transcripts present in a cell is of paramount interest as it reflects the cell's physiological state under the given condition. Genome-wide transcriptomic techniques such as RNA-seq have therefore been used for single-organism analyses for several years, but their applicability has been limited for infection studies.The present work describes the establishment of a novel transcriptomic approach for infection biology which we have termed "Dual RNA-seq". Using this technology, it was intended to shed light particularly on the contribution of non-protein-encoding transcripts to virulence, as these classes have mostly evaded previous infection studies due to the lack of suitable methods. The performance of Dual RNA-seq was evaluated in an in vitro infection model based on the important facultative intracellular pathogen Salmonella enterica serovar Typhimurium and different human cell lines. Dual RNA-seq was found to be capable of capturing all major bacterial and human transcript classes and proved reproducible. During the course of these experiments, a previously largely uncharacterized bacterial small non-coding RNA (sRNA), referred to as STnc440, was identified as one of the most strongly induced genes in intracellular Salmonella.Interestingly, while inhibition of STnc440 expression has been previously shown to cause a virulence defect in different animal models of Salmonellosis, the underlying molecular mechanisms have remained obscure. Here, classical genetics, transcriptomics and biochemical assays proposed a complex model of Salmonella gene expression control that is orchestrated by this sRNA. In particular, STnc440 was found to be involved in the regulation of multiple bacterial target mRNAs by direct base pair interaction with consequences for Salmonella virulence and

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“…T he advent and development of high-throughput sequencing technologies has uncovered the presence of widespread antisense transcription in many bacteria, with the number of annotated genes associated with antisense RNA (asRNA) differing greatly among bacterial species (1,2). asRNAs are encoded on the DNA strand opposite an annotated gene and overlap a portion of a gene or the entire gene, or span multiple genes with perfect complementarity.…”

mentioning

confidence: 99%

“…The mode of regulation by asRNAs can be classified according to molecular mechanism as transcription interference, transcription attenuation, alteration of transcript stability, and translation inhibition (1,2,6). With the exception of transcription interference, a physical RNA-RNA interaction between the sense RNAs and asRNAs is necessary for all of these mechanisms, requiring that both RNAs be expressed in the same cell at the same time.…”

mentioning

confidence: 99%

The double-stranded transcriptome of Escherichia coli

Lybecker

Zimmermann

Bilusic

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

104

146

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Advances in high-throughput transcriptome analyses have revealed hundreds of antisense RNAs (asRNAs) for many bacteria, although few have been characterized, and the number of functional asRNAs remains unknown. We have developed a genomewide high-throughput method to identify functional asRNAs in vivo. Most mechanisms of gene regulation via asRNAs require an RNA-RNA interaction with its target RNA, and we hypothesized that a functional asRNA would be found in a double strand (dsRNA), duplexed with its cognate RNA in a single cell. We developed a method of isolating dsRNAs from total RNA by immunoprecipitation with a ds-RNA specific antibody. Total RNA and immunoprecipitated dsRNA from Escherichia coli RNase III WT and mutant strains were deep-sequenced. A statistical model was applied to filter for biologically relevant dsRNA regions, which were subsequently categorized by location relative to annotated genes. A total of 316 potentially functional asRNAs were identified in the RNase III mutant strain and are encoded primarily opposite to the 5′ ends of transcripts, but are also found opposite ncRNAs, gene junctions, and the 3′ ends. A total of 21 sense/antisense RNA pairs identified in dsRNAs were confirmed by Northern blot analyses. Most of the RNA steady-state levels were higher or detectable only in the RNase III mutant strain. Taken together, our data indicate that a significant amount of dsRNA is formed in the cell, that RNase III degrades or processes these dsRNAs, and that dsRNA plays a major role in gene regulation in E. coli.T he advent and development of high-throughput sequencing technologies has uncovered the presence of widespread antisense transcription in many bacteria, with the number of annotated genes associated with antisense RNA (asRNA) differing greatly among bacterial species (1, 2). asRNAs are encoded on the DNA strand opposite an annotated gene and overlap a portion of a gene or the entire gene, or span multiple genes with perfect complementarity. asRNAs range in size from tens to thousands of nucleotides. Although numerous chromosomally encoded asRNAs have been identified, few have been confirmed by traditional methods or functionally characterized. Raghavan et al. (3) reported that few asRNAs are conserved between Escherichia coli and Salmonella enterica, and that the predicted promoter sequences of the asRNAs are not conserved between these species, suggesting that most asRNA transcripts are products of spurious transcription and are not biologically functional RNAs.The majority of functionally characterized asRNAs are found on plasmids, phages, and transposons (4, 5). The mode of regulation by asRNAs can be classified according to molecular mechanism as transcription interference, transcription attenuation, alteration of transcript stability, and translation inhibition (1, 2, 6). With the exception of transcription interference, a physical RNA-RNA interaction between the sense RNAs and asRNAs is necessary for all of these mechanisms, requiring that both RNAs be expressed in the same...

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Bacterial Antisense RNAs: How Many Are There, and What Are They Doing?

Cited by 324 publications

References 125 publications

An experimentally anchored map of transcriptional start sites in the model cyanobacterium Synechocystis sp. PCC6803

An experimentally anchored map of transcriptional start sites in the model cyanobacterium Synechocystis sp. PCC6803

Dual RNA-seq of pathogen and host

The double-stranded transcriptome of Escherichia coli

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