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Outside a warm-blooded animal host, the enterobacterium Escherichia coli K-12 is also able to grow and survive in stressful nature. The major organic substance in nature is plant, but the genetic system of E. coli how to utilize plant-derived materials as nutrients is poorly understood. Here we describe the set of regulatory targets for uncharacterized IclR-family transcription factor YiaJ on the E. coli genome, using gSELEX screening system. Among a total of 18 high-affinity binding targets of YiaJ, the major regulatory target was identified to be the yiaLMNOPQRS operon for utilization of ascorbate from fruits and galacturonate from plant pectin. The targets of YiaJ also include the genes involved in the utilization for other plant-derived materials as nutrients such as fructose, sorbitol, glycerol and fructoselysine. Detailed in vitro and in vivo analyses suggest that L-ascorbate and α-D-galacturonate are the effector ligands for regulation of YiaJ function. These findings altogether indicate that YiaJ plays a major regulatory role in expression of a set of the genes for the utilization of plant-derived materials as nutrients for survival. PlaR was also suggested to play protecting roles of E. coli under stressful environments in nature, including the formation of biofilm. We then propose renaming YiaJ to PlaR (regulator of plant utilization). The natural hosts of enterobacterium Escherichia coli are warm-blooded animals, but even outside hosts, E. coli can survive even under stressful environments. On earth, the most common organic materials to be used as nutrients by E. coli are plant-derived components, but up to the present time, the genetic system of E. coli for plant utilization is poorly understand. In the course of gSELEX screening of the regulatory targets for hitherto uncharacterized TFs, we identified in this study the involvement of the IclR-family YiaJ in the regulation of about 20 genes or operons, of which the majority are related to the catabolism of plant-derived materials such as ascorbate, galacturonate, sorbitol, fructose and fructoselysine. Therefore, we propose to rename YiaJ to PlaR (regulator of plant utilization). [ABSTRACT FROM AUTHOR]

Escherichia coli contains seven rRNA operons, each consisting of the genes for three rRNAs (16S, 23S and 5S rRNA in this order) and one or two tRNA genes in the spacer between 16S and 23S rRNA genes and one or two tRNA genes in the 3’ proximal region. All of these rRNA and tRNA genes are transcribed from two promoters, P1 and P2, into single large precursors that are afterward processed to individual rRNAs and tRNAs by a set of RNases. In the course of Genomic SELEX screening of promoters recognized by RNA polymerase (RNAP) holoenzyme containing RpoD sigma, a strong binding site was identified within 16S rRNA gene in each of all seven rRNA operons. The binding in vitro of RNAP RpoD holoenzyme to an internal promoter, referred to the promoter of riRNA (an internal RNA of the rRNA operon), within each 16S rRNA gene was confirmed by gel shift assay and AFM observation. Using this riRNA promoter within the rrnD operon as a representative, transcription in vitro was detected with use of the purified RpoD holoenzyme, confirming the presence of a constitutive promoter in this region. LacZ reporter assay indicated that this riRNA promoter is functional in vivo. The location of riRNA promoter in vivo as identified using a set of reporter plasmids agrees well with that identified in vitro. Based on transcription profile in vitro and Northern blot analysis in vivo, the majority of transcript initiated from this riRNA promoter was estimated to terminate near the beginning of 23S rRNA gene, indicating that riRNA leads to produce the spacer-coded tRNA. Under starved conditions, transcription of the rRNA operon is markedly repressed to reduce the intracellular level of ribosomes, but the levels of both riRNA and its processed tRNAGlu stayed unaffected, implying that riRNA plays a role in the continued steady-state synthesis of tRNAs from the spacers of rRNA operons. We then propose that the tRNA genes organized within the spacers of rRNA-tRNA composite operons are expressed independent of rRNA synthesis under specific conditions where further synthesis of ribosomes is not needed. [ABSTRACT FROM AUTHOR]

Enterobacteria are able to survive under stressful conditions within animals, such as acidic conditions in the stomach, bile salts during transfer to the intestine and anaerobic conditions within the intestine. The glutamate-dependent (GAD) system plays a major role in acid resistance in Escherichia coli, and expression of the GAD system is controlled by the regulatory cascade consisting of EvgAS > YdeO > GadE. To understand the YdeO regulon in vivo, we used ChIP-chip to interrogate the E. coli genome for candidate YdeO binding sites. All of the seven operons identified by ChIP-chip as being potentially regulated by YdeO were confirmed as being under the direct control of YdeO using RT-qPCR, EMSA, DNaseI-footprinting and reporter assays. Within this YdeO regulon, we identified four stress-response transcription factors, DctR, NhaR, GadE, and GadW and enzymes for anaerobic respiration. Both GadE and GadW are involved in regulation of the GAD system and NhaR is an activator for the sodium/proton antiporter gene. In conjunction with co-transcribed Slp, DctR is involved in protection against metabolic endoproducts under acidic conditions. Taken all together, we suggest that YdeO is a key regulator of E. coli survival in both acidic and anaerobic conditions. [ABSTRACT FROM AUTHOR]

The transcription factor CsgD plays a key role in the control of biofilm formation in Escherichia coli by controlling the production of curli fimbriae and other biofilm components. In concert with its regulatory role, the promoter for the csgD operon is under the control of more than 10 regulatory factors, each monitoring a different condition or factor in stressful environments. Previously, we classified three factors (OmpR, RstA and IHF) as activators and two factors (CpxR and H-NS) as repressors, and found novel modes of their interplay. Here we describe an as yet uncharacterized regulator, MlrA, that has been suggested to participate in control of curli formation. Based on both in vivo and in vitro analyses, we identified MlrA as a positive factor of the csgD promoter by directly binding to its upstream region (−113 to −146) with a palindromic sequence of AAAATTGTACA(12N)TGTACAATTTT between the binding sites of two activators, IHF and OmpR. The possible interplay between three activators was analysed in detail. [ABSTRACT FROM AUTHOR]

Derivatives of the stationary-phase sigma factor ors of Escherichia coli lacking either of two conserved domains, the postulated N-terminal subregion 1.1 or the C-terminal region 4, were shown to be competent in vitro for transcription initiation from several σ[sup S]-dependent promoters on supercoiled DNA templates. Unlike wild-type ors, however, the deletion derivatives were inactive on relaxed templates. The anomalous slow electrophoretic mobility of σ[sup S] on denaturing gels was corrected by deletion of subregion 1.1, suggesting that this domain in ors may be structurally and functionally analogous to subregion 1.1 of σ[sup 70], substitutions in which have previously been shown to rectify the anomalous electrophoretic migration of σ[sup 70] (V. Gopal and D. Chatterji, Eur. J. Biochem. 244:614-618, 1997). [ABSTRACT FROM AUTHOR]

The article presents a study which analyzes the regulatory roles of MurR (YfeT) transcription factor in both murQP and murR operons of Escherichia coli. It says that the activity of the murP and murR promoter regulators were measured by constructing yfeT (murR)-lacZ and murQ-lacZ protein fusions. The findings reveal that YfeT derepressed the murQP operon of Escherichia coli and the auto-induction of murR was activated by MurR and by other components of the phosphotransferase system.

Transcription profile microarray analysis in Escherichia coli was performed to identify the member genes of the Mg[sup 2+] stimulon that respond to the availability of external Mg[sup 2+] in a PhoP/PhoQ two-component system dependent manner. The mRNA levels of W3110 in the presence of 30 mM MgCl[sub 2], WP3022 (phoP defective), and WQ3007 (phoQ defective) were compared with those of W3110 in the absence of MgCl[sub 2]. The expression ratios of a total of 232 genes were <0.75 in all three strains (the supplemental data are shown at http://www.nara.kindai.ac.jp/nogei/seiken/array.html), suggesting that the PhoP/PhoQ system is involved directly or indirectly in the transcription of these genes. Of those, 26 contained the PhoP box-like sequences with the direct repeats of (T/G)GTTTA within 500 bp upstream of the initiation codon. Furthermore, S1 nuclease assays of 26 promoters were performed to verify six new Mg[sup 2+] stimulon genes, hemL, nagA, rstAB, slyB, vboR, and yrbL, in addition to the phoPQ, mgrB, and mgtA genes reported previously. In gel shift and DNase I footprinting assays, all of these genes were found to be regulated directly by PhoP. Thus, we concluded that the phoPQ, mgrB, mgtA, hemL, nagA, rstAB, slyB, vboR, and yrbL genes make up the Mg[sup 2+] stimulon in E. coli. [ABSTRACT FROM AUTHOR]