Cloning and nucleotide sequence of anaerobically induced porin protein E1 (OprE) of <i>Pseudomonas aeruginosa</i> PAO1

Yamano, Yoshinori; Nishikawa, Takafumi; Komatsu, Yoshihide

doi:10.1111/j.1365-2958.1993.tb01643.x

Cited by 43 publications

(38 citation statements)

References 34 publications

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“…ANR is necessary for denitrification, arginine deiminase activity, and cyanide production by P. aeruginosa under conditions of oxygen limitation (1). That the biofilm cells were deprived of oxygen is supported by the finding that the membrane protein E1 (OprE), an anaerobically induced porin under the control of sigma 54 (55), was also upregulated following attachment. Interestingly, the proteins for the arginine deiminase pathway were found to be downregulated in P. putida 6 h following initial attachment and upregulated again in later stages (47).…”

Section: Discussionmentioning

confidence: 54%

Pseudomonas aeruginosa Displays Multiple Phenotypes during Development as a Biofilm

et al. 2002

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Complementary approaches were employed to characterize transitional episodes in Pseudomonas aeruginosa biofilm development using direct observation and whole-cell protein analysis. Microscopy and in situ reporter gene analysis were used to directly observe changes in biofilm physiology and to act as signposts to standardize protein collection for two-dimensional electrophoretic analysis and protein identification in chemostat and continuous-culture biofilm-grown populations. Using these approaches, we characterized five stages of biofilm development: (i) reversible attachment, (ii) irreversible attachment, (iii) maturation-1, (iv) maturation-2, and (v) dispersion. Biofilm cells were shown to change regulation of motility, alginate production, and quorum sensing during the process of development. The average difference in detectable protein regulation between each of the five stages of development was 35% (approximately 525 proteins). When planktonic cells were compared with maturation-2 stage biofilm cells, more than 800 proteins were shown to have a sixfold or greater change in expression level (over 50% of the proteome). This difference was higher than when planktonic P. aeruginosa were compared with planktonic cultures of Pseudomonas putida. Las quorum sensing was shown to play no role in early biofilm development but was important in later stages. Biofilm cells in the dispersion stage were more similar to planktonic bacteria than to maturation-2 stage bacteria. These results demonstrate that P. aeruginosa displays multiple phenotypes during biofilm development and that knowledge of stage-specific physiology may be important in detecting and controlling biofilm growth.Recent investigations have been directed at determining the degree to which gene regulation during biofilm development controls the switch from planktonic to attached growth. Brözel and coworkers monitored changes in global gene expression patterns in attached Pseudomonas aeruginosa cells and found more than 11 proteins whose levels were altered during various stages of attachment (6). Genevaux and colleagues screened a library of Tn10 insertion mutants of Escherichia coli with altered adhesion abilities (18). Fifty adhesion-deficient mutants were isolated which showed less than 40% attachment compared to the wild type, and 22 mutants were found with an attachment of 40 to 75% compared to the wild type. The majority of these mutants were affected in motility. PrigentCombaret and coworkers (42) carried out a screen in E. coli K-12 similar to that of Genevaux and revealed major changes in the patterns of gene expression during the switch from planktonic to attached growth. They found attachment-dependent regulation of gene expression in 38% of the generated lacZ gene fusions (out of 446 clones). More recently, it has been shown that in Pseudomonas putida, more than 30 genes and 40 gene products were altered within 6 h following attachment (47).These results indicate that physiological changes in the transition from planktonic to attached cells are...

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Section: Discussionmentioning

confidence: 54%

Pseudomonas aeruginosa Displays Multiple Phenotypes during Development as a Biofilm

et al. 2002

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“…The second protein found in increased concentrations in wild-type cells was porin E1 (OprE). The porin protein E1 gene oprE has been described before to be expressed in response to anaerobiosis (Yamano et al, 1993). Furthermore, the two-component class II ribonucleotide reductase subunits NrdJa and NrdJb showed an increased concentration in wild-type cells, suggesting a NarL-dependent induction of the corresponding genes (Torrents et al, 2005).…”

Section: Resultsmentioning

confidence: 86%

Nitrate-responsive NarX-NarL represses arginine-mediated induction of the Pseudomonas aeruginosa arginine fermentation arcDABC operon

et al. 2008

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Denitrification and arginine fermentation are major parts of the anaerobic metabolism of Pseudomonas aeruginosa, which is important for biofilm formation and infection. The twocomponent regulatory system NarX-NarL is part of the underlying network and is required for denitrifying growth. All target promoters identified so far are activated by NarL. In this study the effect of NarL on arginine fermentation was investigated using proteome, Northern blot and lacZ reporter gene analyses. NarL-dependent repression of the arcDABC operon was observed and the corresponding NarL-binding site in the arcD promoter region was functionally localized at "60 bp upstream of the transcriptional start site using site-directed promoter mutagenesis and reporter gene fusion experiments. The results clearly show that in the presence of nitrate NarL represses the arginine-dependent activation of the arcDABC operon mediated by ArgR. It does not influence the oxygen-tension-dependent activation via Anr. Thus, the anaerobic energy metabolism of P. aeruginosa is coordinated via NarX-NarL activity. In the presence of nitrate the highly efficient denitrification is preferred over the less attractive arginine fermentation. INTRODUCTIONIn Pseudomonas aeruginosa anaerobic metabolism is important for biofilm growth and for the infection of the cystic fibrosis lung involving biofilm-like P. aeruginosa microcolonies (Barraud et al., 2006;Filiatrault et al., 2006;Hassett et al., 2002;Palmer et al., 2007;Platt et al., 2008;Sauer et al., 2002;Van Alst et al., 2007;Worlitzsch et al., 2002;Xu et al., 1998). In the absence of oxygen P. aeruginosa grows either by denitrification or by arginine fermentation (Carlson & Ingraham, 1983;Vander Wauven et al., 1984;Zumft, 1997). The expression of genes which encode enzymes for denitrification in P. aeruginosa are controlled by the regulatory proteins Anr and Dnr, as well as the nitrate-responsive two-component system NarXNarL (Arai et al., 1997;Schreiber et al., 2007;Ye et al., 1995). Enzymes required for arginine fermentation are encoded by genes which are organized in the operon arcDABC (Luthi et al., 1990). Anaerobic expression of arcDABC has been shown to be Anr-dependent and further stimulated by the arginine-responsive regulator ArgR (Lu et al., 1999). The global oxygen-sensing regulator Anr is an Escherichia coli Fnr homologue and is essential for fermentative and denitrifying growth in the absence of oxygen (Ye et al., 1995). Moreover, Anr controls the expression of dnr, which encodes the nitrogen-oxidesensing regulator Dnr (Arai et al., 1997). Dnr finally induces expression of the genes encoding the nitrite, nitric oxide and nitrous oxide reductases required for denitrification (Arai et al., 1999(Arai et al., , 2003. Mutant and gene expression studies in P. aeruginosa and Pseudomonas stutzeri showed that the two-component system NarXNarL is additionally required for denitrifying growth (Härtig et al., 1999;Schreiber et al., 2007). The P. aeruginosa NarX protein is a sensor kinase and shares 38 % and 3...

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“…One half of a binding site for Anr was discovered at promoter 1, while promoter 2 lacked any sequence homology. The involvement of one half of the Anr recognition site in oxygen regulation has been proposed for the P. aeruginosa porin gene oprE (70). One explanation for these interesting observations could be provided by the protein-protein interaction of Anr with other, as yet unknown transcription factors involved in hemA expression at the promoter sites found.…”

Section: Resultsmentioning

confidence: 92%

“…A potential half site for the binding of the oxygen regulator Anr, the Pseudomonas Fnr analog, was detected at promoter 1, and it is indicated in Fig. 3 (26,60,70,71). Primer extension experiments were performed by using RNA prepared from aerobically grown P. aeruginosa ( Fig.…”

Section: Resultsmentioning

confidence: 99%

Regulation of the hemA gene during 5-aminolevulinic acid formation in Pseudomonas aeruginosa

et al. 1995

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The general tetrapyrrole precursor 5-aminolevulinic acid is formed in bacteria via two different biosynthetic pathways. Members of the ␣ group of the proteobacteria use 5-aminolevulinic acid synthase for the condensation of succinyl-coenzyme A and glycine, while other bacteria utilize a two-step pathway from aminoacylated tRNA Glu . The tRNA-dependent pathway, involving the enzymes glutamyl-tRNA reductase (encoded by hemA) and glutamate-1-semialdehyde-2,1-aminomutase (encoded by hemL), was demonstrated to be used by Pseudomonas aeruginosa, Pseudomonas putida, Pseudomonas stutzeri, Comamonas testosteroni, Azotobacter vinelandii, and Acinetobacter calcoaceticus. To study the regulation of the pathway, the glutamyl-tRNA reductase gene (hemA) from P. aeruginosa was cloned by complementation of an Escherichia coli hemA mutant. The hemA gene was mapped to the SpeI A fragment and the DpnIL fragment of the P. aeruginosa chromosome corresponding to min 24.1 to 26.8. The cloned hemA gene, coding for a protein of 423 amino acids with a calculated molecular mass of 46,234 Da, forms an operon with the gene for protein release factor 1 (prf1). This translational factor mediates the termination of the protein chain at the ribosome at amber and ochre codons. Since the cloned hemA gene did not possess one of the appropriate stop codons, an autoregulatory mechanism such as that postulated for the enterobacterial system was ruled out. Three open reading frames of unknown function transcribed in the opposite direction to the hemA gene were found. hemM/orf1 and orf2 were found to be homologous to open reading frames located in the 5 region of enterobacterial hemA genes. While orf2 was found to be 59% identical to its enterobacterial counterpart, hemM/orf1 showed only 23% identity. The third open reading frame (orf3), located between the hemA gene and hemM/orf1, encodes a polypeptide with homology to outer membrane proteins. Biochemical and genetic evidence which makes a direct involvement of HemM/ Orf1 in the 5-aminolevulinic acid formation of P. aeruginosa very unlikely was obtained. An increase of hemA mRNA was observed under anaerobic denitrifying conditions, while anaerobic growth utilizing the fermentative arginine deiminase pathway led to a drastic decrease of hemA transcription compared with that observed for aerobically grown P. aeruginosa. These results suggest the anaerobic induction of hemA by the presence of nitrate. Two transcription start sites were located in the 5 region of the hemA gene. Utilization of both transcription start sites was changed in a P. aeruginosa mutant missing the oxygen regulator Anr (Fnr analog), indicating the involvement of the transcription factor in hemA expression. DNA sequences homologous to one half of an Anr binding site were detected at one of the determined transcription start sites.The tetrapyrrole content of a bacterial cell varies drastically depending on the conditions of the growth environment. The formation of 5-aminolevulinic acid (ALA), the first committed precursor of all tetra...

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Cloning and nucleotide sequence of anaerobically induced porin protein E1 (OprE) of Pseudomonas aeruginosa PAO1

Cited by 43 publications

References 34 publications

Pseudomonas aeruginosa Displays Multiple Phenotypes during Development as a Biofilm

Pseudomonas aeruginosa Displays Multiple Phenotypes during Development as a Biofilm

Nitrate-responsive NarX-NarL represses arginine-mediated induction of the Pseudomonas aeruginosa arginine fermentation arcDABC operon

Regulation of the hemA gene during 5-aminolevulinic acid formation in Pseudomonas aeruginosa

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