scholarly journals In vivo genomic footprinting analysis reveals that the complex Bradyrhizobium japonicum fixRnifA promoter region is differently occupied by two distinct RNA polymerase holoenzymes

1998 ◽  
Vol 95 (3) ◽  
pp. 1014-1019 ◽  
Author(s):  
H. Barrios ◽  
R. Grande ◽  
L. Olvera ◽  
E. Morett
Keyword(s):  
Rna Polymerase ◽  
Bradyrhizobium Japonicum ◽  
Promoter Region ◽  
Genomic Footprinting

scholarly journals RNA polymerase II interacts with the promoter region of the noninduced hsp70 gene in Drosophila melanogaster cells.

1986 ◽  
Vol 6 (11) ◽  
pp. 3984-3989 ◽  
Author(s):  
D S Gilmour ◽  
J T Lis
Keyword(s):  
Drosophila Melanogaster ◽  
Heat Shock ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Promoter Region ◽  
High Energy ◽  
Heat Shock Gene ◽  
Cross Linking ◽  
Hsp70 Gene

By using a protein-DNA cross-linking method (D. S. Gilmour and J. T. Lis, Mol. Cell. Biol. 5:2009-2018, 1985), we examined the in vivo distribution of RNA polymerase II on the hsp70 heat shock gene in Drosophila melanogaster Schneider line 2 cells. In heat shock-induced cells, a high level of RNA polymerase II was detected on the entire gene, while in noninduced cells, the RNA polymerase II was confined to the 5' end of the hsp70 gene, predominantly between nucleotides -12 and +65 relative to the start of transcription. This association of RNA polymerase II was apparent whether the cross-linking was performed by a 10-min UV irradiation of chilled cells with mercury vapor lamps or by a 40-microsecond irradiation of cells with a high-energy xenon flash lamp. We hypothesize that RNA polymerase II has access to, and a high affinity for, the promoter region of this gene before induction, and this poised RNA polymerase II may be critical in the mechanism of transcription activation.

scholarly journals Identification and Characterization of the dps Promoter of Mycobacterium smegmatis: Promoter Recognition by Stress-Specific Extracytoplasmic Function Sigma Factors σH and σF

2007 ◽  
Vol 189 (24) ◽  
pp. 8973-8981 ◽  
Author(s):  
Rakhi Pait Chowdhury ◽  
Surbhi Gupta ◽  
Dipankar Chatterji
Keyword(s):  
Rna Polymerase ◽  
Mycobacterium Smegmatis ◽  
Promoter Region ◽  
Sigma Factors ◽  
Galactosidase Activity ◽  
Putative Promoter ◽  
Activity Measurements ◽  
Promoter Recognition

ABSTRACT The survival of a bacterium with a depleted oxygen or nutrient supply is important for its long-term persistence inside the host under stressful conditions. We studied a gene, dps, from Mycobacterium smegmatis, encoding a protein, Dps (for DNA binding protein from starved cells), which is overexpressed under oxidative and nutritional stresses and provides bimodal protection to the bacterial DNA. Characterization of the dps promoter in vivo is therefore important. We cloned a 1-kb putative promoter region of the dps gene of M. smegmatis in an Escherichia coli-Mycobacterium shuttle vector, pSD5B, immediately upstream of the lacZ gene. Promoter activities were assayed in vivo both in solid medium and in liquid cultures by quantitative β-galactosidase activity measurements. To characterize the minimal promoter region, a 200-bp fragment from the whole 1-kb sequence was further cloned in the same vector, and in a similar way, β-galactosidase activity was quantitated. Primer extension analysis was performed to determine the +1 transcription start site of the gene. Point mutations were inserted in the putative promoter sequences in the −10 and −20 regions, and the promoter sequence was confirmed. The promoter was not recognized by purified M. smegmatis core RNA polymerase reconstituted with purified Mycobacterium tuberculosis σA or σB during multiple- and single-round in vitro transcription assays. Promoter-specific in vivo pull-down assays with an immobilized 1-kb DNA fragment containing the dps promoter established that extracellular function sigma factors were associated with this starvation-inducible promoter. Single-round transcription at the dps promoter further supported the idea that only core RNA polymerase reconstituted with σF or σH can generate proper transcripts.

RNA polymerase II interacts with the promoter region of the noninduced hsp70 gene in Drosophila melanogaster cells

1986 ◽  
Vol 6 (11) ◽  
pp. 3984-3989
Author(s):  
D S Gilmour ◽  
J T Lis
Keyword(s):  
Drosophila Melanogaster ◽  
Heat Shock ◽  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Promoter Region ◽  
High Energy ◽  
Heat Shock Gene ◽  
Cross Linking ◽  
Hsp70 Gene

By using a protein-DNA cross-linking method (D. S. Gilmour and J. T. Lis, Mol. Cell. Biol. 5:2009-2018, 1985), we examined the in vivo distribution of RNA polymerase II on the hsp70 heat shock gene in Drosophila melanogaster Schneider line 2 cells. In heat shock-induced cells, a high level of RNA polymerase II was detected on the entire gene, while in noninduced cells, the RNA polymerase II was confined to the 5' end of the hsp70 gene, predominantly between nucleotides -12 and +65 relative to the start of transcription. This association of RNA polymerase II was apparent whether the cross-linking was performed by a 10-min UV irradiation of chilled cells with mercury vapor lamps or by a 40-microsecond irradiation of cells with a high-energy xenon flash lamp. We hypothesize that RNA polymerase II has access to, and a high affinity for, the promoter region of this gene before induction, and this poised RNA polymerase II may be critical in the mechanism of transcription activation.

scholarly journals Promoter Selectivity of the Bradyrhizobium japonicum RpoH Transcription Factors In Vivo and In Vitro

1998 ◽  
Vol 180 (9) ◽  
pp. 2395-2401 ◽  
Author(s):  
Franz Narberhaus ◽  
Michael Kowarik ◽  
Christoph Beck ◽  
Hauke Hennecke
Keyword(s):  
Heat Shock ◽  
Rna Polymerase ◽  
Bradyrhizobium Japonicum ◽  
Normal Growth ◽  
Growth Conditions ◽  
Start Site ◽  
E Coli ◽  
Soluble C

ABSTRACT Expression of the dnaKJ andgroESL 1 heat shock operons ofBradyrhizobium japonicum depends on a ς32-like transcription factor. Three such factors (RpoH1, RpoH2, and RpoH3) have previously been identified in this organism. We report here that they direct transcription from some but not all ς32-type promoters when the respective rpoH genes are expressed inEscherichia coli. All three RpoH factors were purified as soluble C-terminally histidine-tagged proteins, although the bulk of overproduced RpoH3 was insoluble. The purified proteins were recognized by an anti-E. coli ς32 serum. While RpoH1 and RpoH2 productively interacted with E. coli core RNA polymerase and produced E. coli groE transcript in vitro, RpoH3 was unable to do so.B. japonicum core RNA polymerase was prepared and reconstituted with the RpoH proteins. Again, RpoH1 and RpoH2 were active, and they initiated transcription at theB. japonicum groESL 1 and dnaKJpromoters. In all cases, the in vitro start site was shown to be identical to the start site determined in vivo. Promoter competition experiments revealed that the B. japonicum dnaKJ andgroESL 1 promoters were suboptimal for transcription by RpoH1- or RpoH2-containing RNA polymerase from B. japonicum. In a mixture of different templates, the E. coli groESL promoter was preferred over any other promoter. Differences were observed in the specificities of both sigma factors toward B. japonicum rpoH-dependent promoters. We conclude that the primary function of RpoH2is to supply the cell with DnaKJ under normal growth conditions whereas RpoH1 is responsible mainly for increasing the level of GroESL1 after a heat shock.

Sign in / Sign up

Export Citation Format

Share Document