• Korean Journal of Microbiology
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• v.45 no.3
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• pp.263-267
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• 2009

An RpoS factor is a transcriptional regulator which participates in numerous biological processes. In this work, we investigated the transcriptional regulation of proBA and proC composing proline biosynthetic pathway in Escherichia coli. While the proBA and proC genes were greatly induced in an exponential growth phase, they were dramatically repressed in a stationary growth phase in the wild type E. coli. Unlike the wild type E. coli, the proBA and proC genes were not repressed even in the stationary growth phase in its isogenic rpoS mutant. These results suggest that the RpoS factor acts as a transcriptional repressor of proBA and proC genes. The production of threonine, methionine, lysine, and arginine in the rpoS mutant were also increased by more than two times compared to its parental wild type, suggesting that the mutant is able to be used as an useful host strain for the amino acid overproduction.

• Molecules and Cells
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• v.42 no.5
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• pp.426-439
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• 2019

Many small RNAs (sRNAs) regulate gene expression by base pairing to their target messenger RNAs (mRNAs) with the help of Hfq in Escherichia coli. The sRNA DsrA activates translation of the rpoS mRNA in an Hfq-dependent manner, but this activation ability was found to partially bypass Hfq when DsrA is overproduced. The precise mechanism by which DsrA bypasses Hfq is unknown. In this study, we constructed strains lacking all three rpoS-activating sRNAs (i.e., ArcZ, DsrA, and RprA) in $hfq^+$ and $Hfq^-$ backgrounds, and then artificially regulated the cellular DsrA concentration in these strains by controlling its ectopic expression. We then examined how the expression level of rpoS was altered by a change in the concentration of DsrA. We found that the translation and stability of the rpoS mRNA are both enhanced by physiological concentrations of DsrA regardless of Hfq, but that depletion of Hfq causes a rapid degradation of DsrA and thereby decreases rpoS mRNA stability. These results suggest that the observed Hfq dependency of DsrA-mediated rpoS activation mainly results from the destabilization of DsrA in the absence of Hfq, and that DsrA itself contributes to the translational activation and stability of the rpoS mRNA in an Hfq-independent manner.

• Journal of Microbiology and Biotechnology
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• v.14 no.5
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• pp.1063-1066
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• 2004

The rpoS gene product is a global transcriptional factor, which is involved in bacterial survival under various stress conditions. An rpoS-homologous gene was cloned from a septicemia-causing pathogenic Vibrio vulnificus. Introduction of this gene as a multicopy plasmid into various E. coli strains displayed functional complementation, for examples, increased survivability of an rpoS-defective E. coli cell and induction of known $\delta^S$-dependent, stress-responding promoters of E. coli genes.

• Korean Journal of Microbiology
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• v.32 no.4
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• pp.264-270
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• 1994

A gene coding for a novel putative $\sigma$ factor of RNA polymerase has been identified from Streptomyces coelicolor A3(2) using Escherichia coli rpoS gene fragment as a probe. The 486 bp rpoS gene fragment was amplified from E. coli genomic DNA by PCR with two synthetic oligonucleotides, the sequences of which were deduced from the amino acid sequences in the regions 2.3 and 4.2 conserved among various bacterial factors. When E. coli genomic DNA fragments were hybridized with cloned rpoS probe, only one band corresponding to rpoS gene (3.2 kb PvuII fragment or 2.3 kb KpnI fragment) was detected. In S. coelicolor, however, two bands were detected both in PvuII digested DNA and SalI digested DNA. 3.5 kb PvuII fragment which binds the rpoS gene probe was cloned (pMS1) from the sublibrary, and the nucleotide sequences of 1.0 kb BamH'/HincII subclone (pBH2) was partially determined. The nucleotide sequences revealed extensive similarity to other $\sigma$ factor genes of S. coelicolor (hrdA, hrdB, hrdC, hrdD), S. aureofaciens (hrdA, hrdB, hrdC, hrdD), Synechococcus species, Pseudomonas aeruginosa, Stigmatella aurantiaca, and Anabaena species. The nucleotide sequences in regions 1.2 and 4 were compared with the corresponding regions of 5 known ${\sigma}$ factor genes of S. coelicolor by multiple alignment. It turned out that the cloned gene is most closely related to hrdA showing 88% amino acid similarity in region 1.2 and 75% in region 4.

• Proceedings of the Korean Society for Applied Microbiology Conference
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• 2004.06a
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• pp.258-265
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• 2004

Numerous genes of Escherichia coli have been shown to growth phase-dependent expression throughout growth. The global patterns of growth phase-dependent gene expression of E. coli throughout growth using oligonucleotide microarrays containing a nearly complete set of 4,289 annotated open reading frames. To determine the change of gene expression throughout growth, we compared RNAs taken from timecourses with common reference RNA, which is combined with equal amount of RNA pooled from each time point. The hierarchical clustering of the conditions in accordance with timecourse expression revealed that growth phases were clustered into four classes, consistent with known physiological growth status. We analyzed the differences of expression levels at genome level in both exponential and stationary growth phase cultures. Statistical analysis showed that 213 genes are shown to, growth phase-dependent expression. We also analyzed the expression of 256 known operons and 208 regulatory genes. To assess the global impact of RpoS, we identified 193 genes coregulated with rpoS and their expression levels were examined in the isogenic rpoS mutant. The results revealed that 99 of 193 were novel RpoS-dependent stationary phase-induced genes and the majority of those are functionally unknown. Our data provide that global changes and adjustments of gene expression are coordinately regulated by growth transition in E. coli.

• Journal of Microbiology
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• v.35 no.4
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• pp.277-283
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• 1997

Promoters inducible by paraquat, a superocide-generating agent, were isolated from Escherichia coli using a promoter-probing plasmid pRS415 with promoterless lacA gene. Twenty one promoters induced by paraquat were selected and further characterized. From sequence analysis, thirteen of the promoters were mapped to their specific loci on the Escherichia coli chromosome. Several promoters were mapped to the upstream of known genes such as usgl, katG, and mglB, whose relationships with superoxide response have not been previously reported. Other promoters were mapped to the upstream region of unknown open reading frames. Downstream of HC 96 promoter are uncharacterized ORFs whose sequences are homologous to ABC-transporter subunits. Downstream of HC84 promoter is an ORF encoding a transcriptional regulator-like protein, which contains a LysR family-specific HTH (helix-turn-helix) DNA bindign motif. We investigated whether these promoters belong to the soxRS regulon. All promoters except HC96 were found to belong to the soxRS regulon. The HC96 promoter was significantly induced by paraquat in the soxRS deletion mutant strain. The basal transcription level of three promoters (HE43, HC71, HD94) significantly increased at the stationary phase, implying that they are regulated by RpoS. However, paraquat inducibility of all promoters disappeared in the stationary phase, suggesting that SoxRS regulatory system is active only in rapidly growing cells.

• Journal of Microbiology
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• v.45 no.6
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• pp.558-565
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• 2007

Using microarray analysis, we determined those Salmonella genes induced at the entry of stationary phase, and subsequently discovered that uncharacterized yliH was induced most dramatically. We set out to establish the molecular mechanism underlying the stationary phase induction of yliH under the standard culture condition, LB with vigorous aeration, by analyzing its promoter activity in various mutant backgrounds, lacking stationary phase ${\sigma}$, $RpoS^-$, or stringent signal molecules ppGpp, ${\Delta}relA$ ${\Delta}spoT$. It was found that the stationary phase induction of yliHp was partially dependent on rpoS but entirely dependent on ppGpp. DNA sequence analysis revealed that the Salmonella yliH gene is composed of 381 base-pair nucleotides, with overall amino acid sequence revealing 76.38% amino acid identity and 88.98% similarity with Escherichia coli yliH, although no motif from data base was noted for its possible role. Recently however, it has been reported that yliH in E. coli was implicated in biofilm formation and motility by repressing these activities (Domka et al., 2006). We have constructed a mutant Salmonella deleting yliH gene by allele replacement and examined its phenotype, and found that the yliH in Salmonella more or less affects motility and adherence by enhancing these activities. The effect on biofilm formation in Salmonella was uncertain. Moreover, addition of cloned yliH of E. coli into Salmonella did not reduce motility or adherence. Taken together, it appears that the pathways implicating yliH for biofilm formation and motility in E. coli and in Salmonella are somewhat different.

• Journal of Microbiology
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• v.41 no.2
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• pp.109-114
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• 2003

A promoter that is inducible by paraquat and menadione, the superoxide generators, independently of soxRS has been found in front of the sufABCDSE operon in Escherichia coli. Based on the observation that SufA is a holomog of IscA that functions in the assembly of iron sulfur cluster and the sufA promoter (sufAp) contains a putative Fur-binding consensus, we investigated whether this gene is regulated by Fur, a ferric uptake regulator, When examined in several sufAp-lacZ chromosomal fusion strains, sufAp was induced by EDTA, an iron chelator and a well-known Fur-inducer, The basal level of sufA expression increased dramatically in fur mutant, suggesting repression of sufAp by Fur. The derepression in fur mutant and EDTA-induction of sufA expression required nucleotides up to -61, where a putative Fur box is located. Purified Fur protein bound to the DNA fragment containing the putative Fur box between -35 and -10 promoter elements. The regulation by Fur and menadione induction of sufAp acted independently. The rpoS mutation increased sufA induction by menadione, suggesting that the stationary sigma factor RpoS acts negatively on sufA induction.