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  • 1
    Electronic Resource
    Electronic Resource
    An investigation into the compartmentalization of the sporulation transcription factor σE in Bacillus subtilis (2002)
    Oxford, UK : Blackwell Science Ltd.
    Molecular microbiology 43 (2002), S. 0 
    Details
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Sporulation in Bacillus subtilis involves the formation of a polar septum, which divides the sporangium into a mother cell and a forespore. The σE factor, which is encoded within the spoIIG operon, is a cell-specific regulatory protein that directs gene transcription in the mother cell. σE is synthesized as an inactive proprotein pro-σE, which is converted to the mature factor by the putative processing enzyme SpoIIGA. Processing of pro-σE does not commence until after asymmetric division when σE is largely confined to the mother cell. Processing depends on the signalling protein SpoIIR, which delays proteolysis until after polar septation, but the mechanism by which σE is confined to the mother cell is not understood. Previous work favoured a model in which pro-σE localizes to the mother cell face of the polar septum, such that σE would be selectively released into mother cell cytoplasm. Based on the use of green fluorescent protein (GFP) fusions, we now report that pro-σE is distributed approximately uniformly along all membrane surfaces and is not confined to the mother- cell face of the septum. Rather, our results are consistent with a model in which preferential and persistent transcription of the spoIIG operon in the mother cell and degradation of σE in the forespore contribute to the selective accumulation of σE in the mother cell. Persistent transcription of spoIIG after polar septation also contributes to the proper timing of pro-σE processing.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1046/j.1365-2958.2002.02732.x
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  • 2
    Electronic Resource
    Electronic Resource
    Transcription of the principal sigma-factor genes, rpoD and rpoS, in Pseudomonas aeruginosa is controlled according to the growth phase (1994)
    Oxford, UK : Blackwell Publishing Ltd
    Molecular microbiology 13 (1994), S. 0 
    Details
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The rpoS gene encodes the second principal sigma factor of RNA polymerase in stationary-phase cells in Escherichia coli. We examined the transcription of Pseudomonas aeruginosa rpoS as to the growth of ceils. The results of quantitative S1 nuclease mapping of rpoS and rpoD, encoding the principal sigma factor, indicated that the transcription of rpoS is induced in stationary-phase cells, whereas that of rpoD is induced in exponential-phase cells. By high-resolution S1 nuclease mapping, the 5′- and 3′-ends of rpoS mRNA were determined. The results indicated that rpoS is transcribed as a monocistronic mRNA. The sequence preceding the 5 end of rpoS mRNA showed poor homology to the consensus sequences of the previously known promoters. P. aeruginosa rpoS was not transcribed in E coli. By in vitro transcription assaying, P. aeruginosa rpoS was shown to be transcribed by the RNA polymerase fraction containing the principal sigma (σ70) RNA polymerase of P. aeruginosa
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1365-2958.1994.tb00498.x
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  • 3
    Electronic Resource
    Electronic Resource
    ClpC regulates the fate of a sporulation initiation sigma factor, σH protein, in Bacillus subtilis at elevated temperatures (1998)
    Oxford BSL : Blackwell Science Ltd, UK
    Molecular microbiology 29 (1998), S. 0 
    Details
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: Using a strain carrying a clpC–bgaB transcriptional fusion at the amyE locus, we found that the expression of a clpC operon was induced at the end of exponential growth in a σB-independent manner and ceased around T3.5 in the wild type but not in a spo0H mutant. This suggests that some gene product(s) whose expression is dependent on σH function is required for the turn-off of clpC transcription during an early stage of sporulation. A clpC deletion mutant showed a temperature-sensitive sporulation phenotype and exhibited an abnormally large accumulation of σH in the cell at 45°C after T2, at which time the σH level in the wild type had begun to decrease. These results, together with the fact that spo0H transcription in the clpC deletion mutant was similar to that of the wild type, suggested that ClpC may be responsible for the degradation of σH after the accomplishment of its role in sporulation. Moreover, as expected from these results, overproduction of Spo0A was also observed after the initiation of sporulation in the clpC deletion mutant at 45°C.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1046/j.1365-2958.1998.00943.x
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  • 4
    Electronic Resource
    Electronic Resource
    Analysis of HutP-dependent transcription antitermination in the Bacillus subtilis hut operon: identification of HutP binding sites on hut antiterminator RNA and the involvement of the N-terminus of HutP in binding of HutP to the antiterminator RNA (2004)
    Oxford, UK : Blackwell Science Ltd
    Molecular microbiology 51 (2004), S. 0 
    Details
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: We investigated HutP-dependent transcription antitermination of the Bacillus subtilis hut operon. In vitro transcription assays with the B. subtilisσA-containing RNA polymerase indicated that HutP inhibits transcription termination at the internal terminator by binding to the antiterminator on hut mRNA in the presence of histidine. Ethylnitrosourea modification interference assays and mutational analyses of the interference sites showed that interaction of HutP with a region containing three UAG trinucleotide sequences, which is located on top of the antiterminator structure, is critical for hut antitermination in vivo. Results from kinetic analysis of binding of HutP to RNA containing various portions of the antiterminator sequences indicated that secondary structure is required for binding of HutP to the region containing three UAG triplets in the antiterminator. The in vivo HutP antiterminator activity was reduced by the mutations in the N-terminal region of HutP. The HutP variants with H4A, R7A, I9A and Q26A mutations exhibited reduced binding affinities to the antiterminator RNA in vitro. A 25-mer peptide consisting of amino acid residues 2–26 of HutP bound to the antiterminator RNA. These results indicated that the N-terminus of HutP is involved in binding of HutP to the antiterminator RNA.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1046/j.1365-2958.2003.03891.x
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  • 5
    Electronic Resource
    Electronic Resource
    The Spo0A regulon of Bacillus subtilis (2003)
    Oxford, UK : Blackwell Science Ltd
    Molecular microbiology 50 (2003), S. 0 
    Details
    ISSN: 1365-2958
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology , Medicine
    Notes: The master regulator for entry into sporulation in Bacillus subtilis is the DNA-binding protein Spo0A, which has been found to influence, directly or indirectly, the expression of over 500 genes during the early stages of development. To search on a genome-wide basis for genes under the direct control of Spo0A, we used chromatin immunoprecipitation in combination with gene microarray analysis to identify regions of the chromosome at which an activated form of Spo0A binds in vivo. This information in combination with transcriptional profiling using gene microarrays, gel electrophoretic mobility shift assays, using the DNA-binding domain of Spo0A, and bioinformatics enabled us to assign 103 genes to the Spo0A regulon in addition to 18 previously known members. Thus, in total, 121 genes, which are organized as 30 single-gene units and 24 operons, are likely to be under the direct control of Spo0A. Forty of these genes are under the positive control of Spo0A, and 81 are under its negative control. Among newly identified members of the regulon with transcription that was stimulated by Spo0A are genes for metabolic enzymes and genes for efflux pumps. Among  members  with  transcription  that  was in-hibited by Spo0A are genes encoding components of the DNA replication machinery and genes that govern flagellum biosynthesis and chemotaxis. Also in-cluded in the regulon are many (25) genes with products that are direct or indirect regulators of gene transcription. Spo0A is a master regulator for sporulation, but many of its effects on the global pattern of gene transcription are likely to be mediated indirectly by regulatory genes under its control.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1046/j.1365-2958.2003.03818.x
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  • 6
    Electronic Resource
    Electronic Resource
    Transcription analysis of rpoH in Pseudomonas putida (2001)
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology letters 205 (2001), S. 0 
    Details
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: We previously determined the complete DNA sequence of the rpoH gene encoding the heat-shock σ factor (σH) of Pseudomonas putida. In the present study, the transcriptional start sites of rpoH were determined to be 41 nucleotides (T1), 153 nucleotides (T2) and 157 nucleotides (T3) upstream from the translational start codon (AUG) of rpoH by rapid amplification of cDNA 5′-ends. Based on the locations of T2 and T3, a σ70-type promoter (P2) was determined to be located in the open reading frame region of upstream ftsX in addition to the σE-type promoter (P1; DNA Res. 6 (1999) 241). In the in vitro transcription assay with reconstituted RNA polymerases (Eσ70, EσE, EσH and EσS) of Pseudomonas aeruginosa, EσE transcribed rpoH from T1 and Eσ70 transcribed it from T2 and T3. In both cases, the level of transcription was higher at 42°C than at 30°C. No transcript was detected when EσH or EσS was used. These results indicate that EσE and Eσ70 recognize P1 promoter and P2 promoter, respectively, and also prove that the synthesis of rpoH mRNA is inducible upon heat shock.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1574-6968.2001.tb10942.x
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  • 7
    Electronic Resource
    Electronic Resource
    In vitro transcription analysis of rpoD in Pseudomonas aeruginosa PAO1 (1999)
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology letters 180 (1999), S. 0 
    Details
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: The rpoD gene encoding the principal sigma factor (σ70) of Pseudomonas aeruginosa is transcribed from two promoters, PC and PHS. The sequence of PC is similar to the Escherichia coliσ70 consensus promoter sequence and that of PHS is similar to the E. coliσH consensus promoter sequence. Synthesis of rpoD mRNA from PC is constitutive under both steady-state and heat-shock growth conditions, while that of PHS is transiently induced upon heat-shock. To gain a better understanding of the regulation of rpoD expression, we examined in vitro transcription of rpoD using two RNA polymerases (Eσ70 and EσH, containing σ70 and σH, respectively) purified from P. aeruginosa. DNase I footprinting analysis showed specific bindings of Eσ70 and EσH to PC and PHS promoter regions, respectively. In the in vitro runoff transcription assay, EσH transcribed the template from PHS both at 30°C and 42°C but not from PC. However, Eσ70 transcribed rpoD not only from PC both at 30°C and 42°C but also from PHS at 42°C.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1574-6968.1999.tb08811.x
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  • 8
    Electronic Resource
    Electronic Resource
    In vitro transcription system using reconstituted RNA polymerase (Eσ70, EσH, EσE and EσS) of Pseudomonas aeruginosa (2000)
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology letters 183 (2000), S. 0 
    Details
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: We have developed an in vitro transcription system for Pseudomonas aeruginosa genes, using RNA polymerase (RNAP) holoenzyme reconstituted with purified σ protein and RNAP core enzyme. The RNAP core enzyme was directly purified from P. aeruginosa PAO1 cells. The σ factors of P. aeruginosa (σ70, σH, σE and σS) were prepared in a hexa-histidine tagged form, which were expressed in Escherichia coli and purified using a HisTrap Chelating column. The RNAP holoenzyme reconstituted from core enzyme with each σ factor recognized correctly each of the cognate promoters. This system will be useful for the promoter analysis of many genes in P. aeruginosa.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1574-6968.2000.tb08967.x
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  • 9
    Electronic Resource
    Electronic Resource
    Transcription of the groESL operon in Pseudomonas aeruginosa PAO1 (1998)
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology letters 163 (1998), S. 0 
    Details
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Northern hybridization and S1 nuclease mapping demonstrated that the groES and groEL genes in Pseudomonas aeruginosa were transcribed as a bicistronic mRNA of 2.2 kb. Two transcription start sites and a transcription termination site were mapped. Overlapping consensus sequences for σ32- and σ70-dependent promoters were found in the upstream region of groES. Levels of groESL-specific mRNA were increased about 2-fold upon heat shock. This response differs from the dramatic enhancement (more than 10-fold) of groESL transcription after heat shock observed in other bacterial species.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1574-6968.1998.tb13051.x
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  • 10
    Electronic Resource
    Electronic Resource
    Heterologous expression of the cytochrome P450cam hydroxylase operon and the repressor gene of Pseudomonas putida in Escherichia coli (1994)
    Oxford, UK : Blackwell Publishing Ltd
    FEMS microbiology letters 123 (1994), S. 0 
    Details
    ISSN: 1574-6968
    Source: Blackwell Publishing Journal Backfiles 1879-2005
    Topics: Biology
    Notes: Abstract The cytochrome P450cam hydroxylase operon (camDCAB) of Pseudomonas putida is negatively regulated by a repressor, CamR, which also represses its own gene. The expression in P. putida of both camR and camDCAB is depressed in the presence of d-camphor. We examined the expression in Escherichia coli of camR and camDCAB by monitoring the enzyme activity of the camD gene product. In the presence or absence of d-camphor in the cell culture, the expression in E. coli of camD was significant and constitutive, suggesting no expression of camR. This lack of expression was confirmed by monitoring the β-galactosidase activity of camR-lacZ translational fusions. However, S1 nuclease mapping revealed that synthesis of camR mRNA in E. coli was significant and constitutive, as observed in the case of camDCAB mRNA. Thus, it is likely that the expression of camR in E. coli is limited at the translational level.
    Type of Medium: Electronic Resource
    URL: http://dx.doi.org/10.1111/j.1574-6968.1994.tb07200.x
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