Phenotypic characterization and complementation analysis of Bacillus subtilis 6S RNA single and double deletion mutants

Hoch, Philipp G.; Burenina, Olga Y.; Weber, Michaël; Elkina, Daria A.; Nesterchuk, Mikhail V.; Сергиев, Петр В.; Hartmann, Roland K.; Кубарева, Е. А.

doi:10.1016/j.biochi.2014.12.019

Cited by 26 publications

(39 citation statements)

References 40 publications

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“…We observed that deletion of the 6S-2 RNA gene leads to an up-regulation of numerous proteins involved in metabolism and stress responses, and for exponentially growing cells, this effect was stronger for a 6S-2 than a 6S-1 RNA deletion (Hoch et al 2015). Several of the identified proteins were also upregulated in the 6S-1 RNA deletion strain (in stationary phase), suggesting that there is functional overlap between the two 6S RNAs.…”

Section: Discussionmentioning

confidence: 82%

“…While a 6S-2 RNA deletion resulted in no measurable growth phenotype under standard laboratory conditions (Cavanagh et al 2012), or only in a minor phenotype under alkaline stress conditions (Hoch et al 2015), this does not necessarily imply that 6S-2 RNA is functionally silent. We observed that deletion of the 6S-2 RNA gene leads to an up-regulation of numerous proteins involved in metabolism and stress responses, and for exponentially growing cells, this effect was stronger for a 6S-2 than a 6S-1 RNA deletion (Hoch et al 2015).…”

Section: Discussionmentioning

confidence: 96%

“…Based on Northern blots using T7 transcripts of 6S RNAs as quantitative standards, we recently estimated 6S-2 RNA levels to be roughly threefold higher than 6S-1 RNA levels during exponential phase (OD 600 ∼0.9; Hoch et al 2015). In view of the RNA-seq data (Table 1) suggesting roughly equal numbers for 6S-1 and 6S-2 pRNAs for wt cells during exponential phase, this may be taken as evidence that σ A -RNAP less actively synthesizes pRNAs on 6S-2 RNA compared with 6S-1 RNA.…”

Section: Discussionmentioning

confidence: 99%

“…Surprisingly, a minority of bacteria, mainly those belonging to the firmicutes but also some others like the γ-proteobacterium Legionella pneumophila, express at least two different 6S RNAs (for review, see Wehner et al 2014;Burenina et al 2015). In the case of B. subtilis-the major model system for the study of bacteria expressing two 6S RNA paralogs-the available data suggest differential as well as overlapping roles of 6S-1 and 6S-2 RNAs (Cavanagh et al 2012Cabrera-Ostertag et al 2013;Hoch et al 2015). Further investigations are required to better understand the biological reasons why a second 6S RNA paralog with a different expression profile has evolved in B. subtilis and related bacteria.…”

Section: Discussionmentioning

confidence: 99%

“…ΔbsrAB (ΔbsrA/ΔbsrB, deletion of both 6S RNA genes), ΔbsrAB+A (bsrA reintegrated into the ΔbsrAB strain at the amyE locus under control of the native 6S-1 RNA gene promoter), and ΔbsrAB+B (bsrB reintegrated into the ΔbsrAB strain at the amyE locus under control of the native 6S-2 RNA gene promoter) (Hoch et al 2015). 6S-2 RNA-derived pRNA reads were exclusively detected in strains expressing 6S-2 RNA (wt, ΔbsrA and ΔbsrAB+B), but not in those with a deletion of the bsrB gene (ΔbsrB and ΔbsrAB+A; Table 1; Fig.…”

Section: Smentioning

confidence: 99%

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Bacillus subtilis 6S-2 RNA serves as a template for short transcripts in vivo

Hoch

Schlereth

Lechner

et al. 2016

RNA

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The global transcriptional regulator 6S RNA is abundant in a broad range of bacteria. The RNA competes with DNA promoters for binding to the housekeeping RNA polymerase (RNAP) holoenzyme. When bound to RNAP, 6S RNA serves as a transcription template for RNAP in an RNA-dependent RNA polymerization reaction. The resulting short RNA transcripts (so-called product RNAs = pRNAs) can induce a stable structural rearrangement of 6S RNA when reaching a certain length. This rearrangement leads to the release of RNAP and thus the recovery of transcription at DNA promoters. While most bacteria express a single 6S RNA, some harbor a second 6S RNA homolog (termed 6S-2 RNA in Bacillus subtilis). Bacillus subtilis 6S-2 RNA was recently shown to exhibit essentially all hallmark features of a bona fide 6S RNA in vitro, but evidence for the synthesis of 6S-2 RNAderived pRNAs in vivo has been lacking so far. This raised the question of whether the block of RNAP by 6S-2 RNA might be lifted by a mechanism other than pRNA synthesis. However, here we demonstrate that 6S-2 RNA is able to serve as a template for pRNA synthesis in vivo. We verify this finding by using three independent approaches including a novel primer extension assay. Thus, we demonstrate the first example of an organism that expresses two distinct 6S RNAs that both exhibit all mechanistic features defined for this type of regulatory RNA.

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Section: Discussionmentioning

confidence: 82%