A dual function for a bacterial small RNA: SgrS performs base pairing-dependent regulation and encodes a functional polypeptide

201

240

Regulation of bacterial iron homeostasis is often controlled by the iron-sensing ferric uptake repressor (Fur). The Bacillus subtilis Fur protein acts as an iron-dependent repressor for siderophore biosynthesis and iron transport proteins. Here, we demonstrate that Fur also coordinates an iron-sparing response that acts to repress the expression of iron-rich proteins when iron is limiting. When Fur is inactive, numerous iron-containing proteins are downregulated, including succinate dehydrogenase, aconitase, cytochromes, and biosynthetic enzymes for heme, cysteine, and branched chain amino acids. As a result, a fur mutant grows slowly in a variety of nutrient conditions. Depending on the growth medium, rapid growth can be restored by mutations in one or more of the molecular effectors of the iron-sparing response. These effectors include the products of three Fur-regulated operons that encode a small RNA (FsrA) and three small, basic proteins (FbpA, FbpB, and FbpC). Extensive complementarity between FsrA and the leader region of the succinate dehydrogenase operon is consistent with an RNA-mediated translational repression mechanism for this target. Thus, iron deprivation in B. subtilis activates pathways to remodel the proteome to preserve iron for the most critical cellular functions.

Section: Resultsmentioning

confidence: 99%

The Bacillus subtilis iron-sparing response is mediated by a Fur-regulated small RNA and three small, basic proteins

Gaballa

Antelmann

Aguilar

et al. 2008

201

240

“…1A). The minimal base-pairing region of SgrS is located at nucleotides 168-181 in the 3′ portion (11), whereas the 5′ portion of SgrS encodes a polypeptide of 43 amino acids designated SgrT (12). As the first step to define the functional Hfq-binding site on SgrS, we deleted nucleotides 1-167 of the sgrS gene under the arabinose-inducible promoter on the low copy number plasmid (Fig.…”

Section: Resultsmentioning

confidence: 99%

PolyU tail of rho-independent terminator of bacterial small RNAs is essential for Hfq action

Otaka

Ishikawa

Morita

et al. 2011

172

214

Major bacterial small RNAs (sRNAs) regulate the translation and stability of target mRNAs through base pairing with the help of the RNA chaperone Hfq. The Hfq-dependent sRNAs consist of three basic elements, mRNA base-pairing region, Hfq-binding site, and rho-independent terminator. Although the base-pairing region and the terminator are well documented in many sRNAs, the Hfq-binding site is less well-defined except that Hfq binds RNA with a preference for AU-rich sequences. Here, we performed mutational and biochemical studies to define the sRNA site required for Hfq action using SgrS as a model sRNA. We found that shortening terminator polyU tail eliminates the ability of SgrS to bind to Hfq and to silence ptsG mRNA. We also demonstrate that the SgrS terminator can be replaced with any foreign rho-independent terminators possessing a polyU tail longer than 8 without losing the ability to silence ptsG mRNA in an Hfq-dependent manner. Moreover, we found that shortening the terminator polyU tail of several other sRNAs also eliminates the ability to bind to Hfq and to regulate target mRNAs. We conclude that the polyU tail of sRNAs is essential for Hfq action in general. The data also indicate that the terminator polyU tail plays a role in Hfq-dependent stabilization of sRNAs.riboregulation | RNA 3′ end | RNase E | RyhB

“…Moreover, the 5Ј region of the Ϸ220-nt E. coli SgrS RNA encodes a functional peptide, whereas the 3Ј region contains a regulatory RNA domain that targets ptsG mRNA by base pairing (47). Besides such potential dual functions, some of the short ORFs identified in M. mazei were associated with a cis-encoded asRNA overlapping the 5Ј or 3Ј region of the short ORF (Fig.…”

mentioning

confidence: 99%

Deep sequencing analysis of the Methanosarcina mazei Gö1 transcriptome in response to nitrogen availability

Jäger

Sharma

Thomsen³

et al. 2009

190

243

Methanosarcina mazei and related mesophilic archaea are the only organisms fermenting acetate, methylamines, and methanol to methane and carbon dioxide, contributing significantly to greenhouse gas production. The biochemistry of these metabolic processes is well studied, and genome sequences are available, yet little is known about the overall transcriptional organization and the noncoding regions representing 25% of the 4.01-Mb genome of M. mazei. We present a genome-wide analysis of transcription start sites (TSS) in M. mazei grown under different nitrogen availabilities. Pyrosequencing-based differential analysis of primary vs. processed 5 ends of transcripts discovered 876 TSS across the M. mazei genome. Unlike in other archaea, in which leaderless mRNAs are prevalent, the majority of the detected mRNAs in M. mazei carry long untranslated 5 regions. Our experimental data predict a total of 208 small RNA (sRNA) candidates, mostly from intergenic regions but also antisense to 5 and 3 regions of mRNAs. In addition, 40 new small mRNAs with ORFs of <30 aa were identified, some of which might have dual functions as mRNA and regulatory sRNA. We confirmed differential expression of several sRNA genes in response to nitrogen availability. Inspection of their promoter regions revealed a unique conserved sequence motif associated with nitrogen-responsive regulation, which might serve as a regulator binding site upstream of the common IIB recognition element. Strikingly, several sRNAs antisense to mRNAs encoding transposases indicate nitrogen-dependent transposition events. This global TSS map in archaea will facilitate a better understanding of transcriptional and posttranscriptional control in the third domain of life.Methanosarcina mazei strain Gö1 is a representative methaneproducing archaeon of ecologic significance because of its role in biogenic methane production in various anaerobic habitats on Earth (1). The genome sequences of M. mazei and its close relatives Methanosarcina acetivorans and Methanosarcina barkeri have recently become available and have revealed an unexpected low proportion of coding region (74.2% in M. acetivorans, 75.15% in M. mazei, and 79.2% in M. barkeri) (2-4). The biochemical basis of methanogenesis has been analyzed in considerable detail (5, 6). In contrast, little is known about global regulatory networks that ensure survival in periods of nutrient starvation or stress in this important group of archaea. More than 50 predicted transcriptional regulators were annotated in the genome of M. mazei. Strikingly, most of them seem to be closely related to bacterial proteins (2), whereas the basic components of the archaeal transcription and translation machineries generally are more similar to those of eukaryotes (7). A recent genetic study (8) discovered the first global transcriptional regulator of M. mazei, the nitrogen regulator NrpR, which was experimentally demonstrated to globally repress transcription of nitrogen fixation and assimilation genes in response to the nitrogen source.Bes...