• The Plant Pathology Journal
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• 제29권3호
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• pp.323-329
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• 2013

The stationary-phase sigma factor, RpoS, influences the expression of factors important in survival of Pseudomonas chlororaphis O6 in the rhizosphere. A partial proteomic profile of a rpoS mutant in P. chlororaphis O6 was conducted to identify proteins under RpoS regulation. Five of 14 differentially regulated proteins had unknown roles. Changes in levels of proteins in P. chlororaphis O6 rpoS mutant were associated with iron metabolism, and protection against oxidative stress. The P. chlororaphis O6 rpoS mutant showed increased production of a pyoverdine-like siderophore, indole acetic acid, and altered isozyme patterns for peroxidase, catalase and superoxide dismutase. Consequently, sensitivity to hydrogen peroxide exposure increased in the P. chlororaphis O6 rpoS mutant, compared with the wild type. Taken together, RpoS exerted regulatory control over factors important for the habitat of P. chlororaphis O6 in soil and on root surfaces. The properties of several of the proteins in the RpoS regulon are currently unknown.

• 미생물학회지
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• 제45권3호
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• pp.263-267
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• 2009

세포정지기 및 스트레스에서 유도되는 RpoS 인자는 다양한 세포반응에 관여하는 유전자의 전사발현에 관여하는 전자조절인자이다. 대장균에서 proline의 생합성에 관여하는 유전자인 proBA와 proC 발현을 세포생장 주기별로 조사해 본 결과, 세포지수기에서는 proBA와 proC 유전자의 발현이 유도되는데 비해, 세포정지기에서는 이 세유전자의 전사발현이 극적으로 저해되었다. 그러나 rpoS 돌연변이를 야생형 대장균에 도입한 결과 proline 생합성에 관여하는 유전자인 proBA와 proC 유전자의 발현이 세포정지기에서 저해되지 않았다. 이러한 결과는 RpoS가 proline 생합성에 관여하는 proBA와 proC 유전자의 전사발현에 음성효과를 미치고 있음을 의미한다. 한편 rpoS 돌연변이 균주에서는 proline 외에도 threonine, methionine, lysine, arginine 등의 아미노산이 야생형 대비 2배 이상 생합성이 증가되었는데, 이는 rpoS 대장균 돌연변이 균주가 아미노산의 대량생산에 이용될 수 있음을 의미한다.

• 식물병연구
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• 제30권2호
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• pp.148-156
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• 2024

Ralstonia pseudosolanacearum은 토양과 물에서 오랫동안 생존하고, 가지과 작물에 심각한 풋마름병을 일으키는 식물병원세균이다. Simga S는 세균의 스트레스 환경에서 반응 또는 정지기 동안 유전자 발현을 조절하는 RNA 중합효소 복합체의 일부인 단백질이다. 본 연구는 스트레스 조건에서 R.pseudosolanacearum의 sigma S의 역할을 조사하기 위해서, R.pseudosolanacearum의 GMI1000 균주의 sigma S를 암호화하는 rpoS 유전자 변이체를 준비하여 야생형 균주와 세균의 특징을 비교하였다. 아울러 rpoS 유전자 역할은 원래 유전자를 변이체에 도입하여 rpoS 유전자 표현형 회복을 확인하였다. 야생형 균주와 rpoS 결여 변이체는 생장 속도, 외피다당류 생산, 식물체에서 병원성, 식물 세포벽 분해 효소 활성에서 차이를 보이지 않았다. 그러나 야생형 균주는 영양분결핍 조건에서 변이체보다 더 민감하게 반응하였고 과산화수소가 첨가된 조건에서 변이체보다 덜 민감하게 반응하였다. 흥미롭게도 영양분결핍 조건에서 rpoS 결여 변이체에서는 장기간 생균수를 유지하지만, 같은 조건에서 야생형 균주 생균수는 빠르게 감소하였다. 그리고 두 균주 배양액 pH를 측정한 결과, 야생형 균주와 변이체 간에 상당한 차이가 나타났다. 야생형 균주는 생장하면서 빠르게 배지의 pH가 감소하여 산성화되었다. 그러므로 야생형 균주의 빠른 사멸은 배지가 산성화되면서 정지기 상태 세균의 산성 pH에 대한 민감도 때문일 것이다. Biolog 분석으로 rpoS 변이체는 acetic acid, D-alanine, D-trehalose, L-histidine을 이용하지 못함을 확인하였다. 본 연구 결과는 R. pseudosolanacearum 세균의 sigma S가 영양분결핍 조건에서 정지기 동안 유기산 생산 또는 이용을 조절하며 정지기 세포사멸도 조절하는 것을 보여준다.

• Journal of Microbiology and Biotechnology
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• 제29권8호
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• pp.1299-1309
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• 2019

As an opportunistic bacterial pathogen, Pseudomonas aeruginosa PAO1 contains two phenazine-producing gene operons, phzA1B1C1D1E1F1G1 (phz1) and phzA2B2C2D2E2F2G2 (phz2), each of which is independently capable of encoding all enzymes for biosynthesizing phenazines, including phenazine-1-carboxylic acid and its derivatives. Other previous study reported that the RpoS-deficient mutant SS24 overproduced pyocyanin, a derivative of phenazine-1-carboxylic acid. However, it is not known how RpoS mediates the expression of two phz operons and regulates pyocyanin biosynthesis in detail. In this study, with deletion of the rpoS gene in the $PA{\Delta}phz1$ mutant and the $PA{\Delta}phz2$ mutant respectively, we demonstrated that RpoS exerted opposite regulatory roles on the expression of the phz1and phz2 operons. We also confirmed that the phz1 operon played a critical role and especially biosynthesized much more phenazines than the phz2 operon when the rpoS gene was knocked out in P. aeruginosa. By constructing the translational reporter fusion vector lasR'-'lacZ and the chromosomal fusion mutant $PA{\Delta}lasR::lacZ$, we verified that RpoS deficiency caused increased expression of lasR, a transcription regulator gene in a first quorum sensing system (las) that activates overexpression of the phz1 operon, suggesting that in the absence of RpoS, LasR might act as an intermediate in overproduction of phenazine biosynthesis mediated by the phz1 operon in P. aeruginosa.

• 한국식물병리학회:학술대회논문집
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• 한국식물병리학회 2003년도 정기총회 및 추계학술발표회
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• pp.100.2-101
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• 2003

The global regulator, GacS (global antibiotic and cyanide sensor kinase), was required for the increased resistance to hydrogen peroxide occurring as cultures of the rhizobacterium, P. chlororaphis O6, matured. Specific stationary-phase peroxidase and catalase isozymes were absent in the GacS mutant, whereas a manganese-superoxide dismutase isozyme was expressed earlier and to a great extent than wild type. In the wild type cell, transcript accumulation of rpoS was higher in late logarithmic-phase cells than cells from mid logarithmic- or stationary-phase. Transcripts from rpoS in the GacS mutant were reduced in each of these growth phases compared to the wild type expression. The down stream sequence from rpoS lacked sequences encoding a small RNA, rsmZ, found in other pseudomonads and implicated in control of genes activated by the GacS system. These findings suggest that GacS-mediated regulation of RpoS plays role in control of oxidative stress in P. chlororaphis O6 by as yet an unknown mechanism.

• Journal of Microbiology and Biotechnology
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• 제19권12호
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• pp.1527-1535
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• 2009

Polyphosphate (polyP) plays diverse physiological functions in prokaryotes and eukaryotes, but most of their detailed mechanisms are still obscure. Here, we show that deletion of polyphosphate kinase (PPK), the principal enzyme responsible for synthesis of polyP, resulted in augmented expression of cAMP receptor protein (CRP) and rpoS and lowered $H_2O_2$ sensitivity in Salmonella Typhimurium ATCC14028. The binding of cAMP-CRP complex to rpoS promoter and further stimulation of its transcription were proved through electrophoretic mobility shift assay, lacZ fusion, and exogenous cAMP addition, respectively. The rpoS expression increased in cpdA (cAMP phosphodiesterase coding gene) mutant, further suggesting that cAMP-CRP upregulated rpoS expression. These results demonstrate that PPK affects oxidative stress response by modulating crp and rpoS expression in S. Typhimurium.

• Genomics & Informatics
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• 제12권4호
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• pp.276-282
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• 2014

The disease tuberculosis, caused by Mycobacterium tuberculosis (MTB), remains a major cause of morbidity and mortality in developing countries. The evolution of drug-resistant tuberculosis causes a foremost threat to global health. Most drug-resistant MTB clinical strains are showing resistance to isoniazid and rifampicin (RIF), the frontline anti-tuberculosis drugs. Mutation in rpoB, the beta subunit of DNA-directed RNA polymerase of MTB, is reported to be a major cause of RIF resistance. Amongst mutations in the well-defined 81-base-pair central region of the rpoB gene, mutation at codon 450 (S450L) and 445 (H445Y) is mainly associated with RIF resistance. In this study, we modeled two resistant mutants of rpoB (S450L and H445Y) using Modeller9v10 and performed a docking analysis with RIF using AutoDock4.2 and compared the docking results of these mutants with the wild-type rpoB. The docking results revealed that RIF more effectively inhibited the wild-type rpoB with low binding energy than rpoB mutants. The rpoB mutants interacted with RIF with positive binding energy, revealing the incapableness of RIF inhibition and thus showing resistance. Subsequently, this was verified by molecular dynamics simulations. This in silico evidence may help us understand RIF resistance in rpoB mutant strains.

• 한국미생물생명공학회:학술대회논문집
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• 한국미생물생명공학회 2004년도 Annual Meeting BioExibition International Symposium
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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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• 제41권2호
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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.

• 한국정보통신학회논문지
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• 제17권4호
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• pp.1005-1009
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• 2013

마산병원과 결핵연구원에서 rifampin 내성균주를 확보하여 기존의 전통방법으로 검사된 항결핵제 내성균의 rifampin 내성관련 유전자인 rpoB (RNA polymerase beta subunit)의 변이를 DNA 염기서열 분석방법에 의하여 분석하였다. 그 결과 국내에서 분리되는 결핵균에서는 기존에 보고된 rpoB 변이부위와는 다른 위치의 DNA 염기 변이가 확인되었고 국내에서 여러 내성 결핵균주에서 변이가 발견되고 있지만 이러한 변이가 리팜핀 내성을 실제로 유발하는지 실험적으로 검증된 바는 없었다. 따라서 이러한 특이 부위의 rpoB변이들이 실제로 리팜핀 내성을 유발하는가를 확인하기 위하여 내성 결핵균주들의 rpoB 변이유전자를 polymerase chain reaction(PCR)으로 증폭하고 이것을 리팜핀 감수성 결핵균주에 cloning(클로닝)하고 발현시켜 rpoB 변이가 리팜핀에 대한 내성을 발생시켰음을 실험적으로 확인하였다.