TY - JOUR A1 - Fröhlich, Kathrin S. A1 - Haneke, Katharina A1 - Papenfort, Kai A1 - Vogel, Jörg T1 - The target spectrum of SdsR small RNA in Salmonella JF - Nucleic Acids Research N2 - Model enteric bacteria such as Escherichia coli and Salmonella enterica express hundreds of small non-coding RNAs (sRNAs), targets for most of which are yet unknown. Some sRNAs are remarkably well conserved, indicating that they serve cellular functions that go beyond the necessities of a single species. One of these ‘core sRNAs’ of largely unknown function is the abundant ∼100-nucleotide SdsR sRNA which is transcribed by the general stress σ-factor, σ\(^{S}\) and accumulates in stationary phase. In Salmonella, SdsR was known to inhibit the synthesis of the species-specific porin, OmpD. However, sdsR genes are present in almost all enterobacterial genomes, suggesting that additional, conserved targets of this sRNA must exist. Here, we have combined SdsR pulse-expression with whole genome transcriptomics to discover 20 previously unknown candidate targets of SdsR which include mRNAs coding for physiologically important regulators such as the carbon utilization regulator, CRP, the nucleoid-associated chaperone, StpA and the antibiotic resistance transporter, TolC. Processing of SdsR by RNase E results in two cellular SdsR variants with distinct target spectra. While the overall physiological role of this orphan core sRNA remains to be fully understood, the new SdsR targets present valuable leads to determine sRNA functions in resting bacteria. KW - sRNA KW - Salmonella enterica KW - SdsR Y1 - 2016 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-175365 VL - 44 IS - 21 ER - TY - JOUR A1 - Babski, Julia A1 - Haas, Karina A. A1 - Näther-Schindler, Daniela A1 - Pfeiffer, Friedhelm A1 - Förstner, Konrad U. A1 - Hammelmann, Matthias A1 - Hilker, Rolf A1 - Becker, Anke A1 - Sharma, Cynthia M. A1 - Marchfelder, Anita A1 - Soppa, Jörg T1 - Genome-wide identification of transcriptional start sites in the haloarchaeon Haloferax volcanii based on differential RNA-Seq (dRNA-Seq) JF - BMC Genomics N2 - Background Differential RNA-Seq (dRNA-Seq) is a recently developed method of performing primary transcriptome analyses that allows for the genome-wide mapping of transcriptional start sites (TSSs) and the identification of novel transcripts. Although the transcriptomes of diverse bacterial species have been characterized by dRNA-Seq, the transcriptome analysis of archaeal species is still rather limited. Therefore, we used dRNA-Seq to characterize the primary transcriptome of the model archaeon Haloferax volcanii. Results Three independent cultures of Hfx. volcanii grown under optimal conditions to the mid-exponential growth phase were used to determine the primary transcriptome and map the 5′-ends of the transcripts. In total, 4749 potential TSSs were detected. A position weight matrix (PWM) was derived for the promoter predictions, and the results showed that 64 % of the TSSs were preceded by stringent or relaxed basal promoters. Of the identified TSSs, 1851 belonged to protein-coding genes. Thus, fewer than half (46 %) of the 4040 protein-coding genes were expressed under optimal growth conditions. Seventy-two percent of all protein-coding transcripts were leaderless, which emphasized that this pathway is the major pathway for translation initiation in haloarchaea. A total of 2898 of the TSSs belonged to potential non-coding RNAs, which accounted for an unexpectedly high fraction (61 %) of all transcripts. Most of the non-coding TSSs had not been previously described (2792) and represented novel sequences (59 % of all TSSs). A large fraction of the potential novel non-coding transcripts were cis-antisense RNAs (1244 aTSSs). A strong negative correlation between the levels of antisense transcripts and cognate sense mRNAs was found, which suggested that the negative regulation of gene expression via antisense RNAs may play an important role in haloarchaea. The other types of novel non-coding transcripts corresponded to internal transcripts overlapping with mRNAs (1153 iTSSs) and intergenic small RNA (sRNA) candidates (395 TSSs). Conclusion This study provides a comprehensive map of the primary transcriptome of Hfx. volcanii grown under optimal conditions. Fewer than half of all protein-coding genes have been transcribed under these conditions. Unexpectedly, more than half of the detected TSSs belonged to several classes of non-coding RNAs. Thus, RNA-based regulation appears to play a more important role in haloarchaea than previously anticipated. KW - Archaea KW - dRNA-Seq KW - Promoter KW - Non-coding RNAs KW - sRNA KW - Haloferax volcanii KW - Transcriptome KW - Leaderless transcript KW - Antisense RNA Y1 - 2016 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-164553 VL - 17 IS - 629 ER - TY - JOUR A1 - Hershko-Shalev, Tal A1 - Odenheimer-Bergman, Ahuva A1 - Elgrably-Weiss, Maya A1 - Ben-Zvi, Tamar A1 - Govindarajan, Sutharsan A1 - Seri, Hemda A1 - Papenfort, Kai A1 - Vogel, Jörg A1 - Altuvia, Shoshy T1 - Gifsy-1 Prophage IsrK with Dual Function as Small and Messenger RNA Modulates Vital Bacterial Machineries JF - PLoS Genetics N2 - While an increasing number of conserved small regulatory RNAs (sRNAs) are known to function in general bacterial physiology, the roles and modes of action of sRNAs from horizontally acquired genomic regions remain little understood. The IsrK sRNA of Gifsy-1 prophage of Salmonella belongs to the latter class. This regulatory RNA exists in two isoforms. The first forms, when a portion of transcripts originating from isrK promoter reads-through the IsrK transcription-terminator producing a translationally inactive mRNA target. Acting in trans, the second isoform, short IsrK RNA, binds the inactive transcript rendering it translationally active. By switching on translation of the first isoform, short IsrK indirectly activates the production of AntQ, an antiterminator protein located upstream of isrK. Expression of antQ globally interferes with transcription termination resulting in bacterial growth arrest and ultimately cell death. Escherichia coli and Salmonella cells expressing AntQ display condensed chromatin morphology and localization of UvrD to the nucleoid. The toxic phenotype of AntQ can be rescued by co-expression of the transcription termination factor, Rho, or RNase H, which protects genomic DNA from breaks by resolving R-loops. We propose that AntQ causes conflicts between transcription and replication machineries and thus promotes DNA damage. The isrK locus represents a unique example of an island-encoded sRNA that exerts a highly complex regulatory mechanism to tune the expression of a toxic protein. KW - prophage KW - Gifsy-1 KW - sRNA KW - IsrK Y1 - 2016 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-166717 VL - 12 IS - 4 ER -