DOI: 10.1101/gad.7.1.149. Role of the sigma 70 subunit of RNA polymerase in transcriptional activation by activator protein PhoB in Escherichia coli.

Role of the sigma 70 subunit of RNA polymerase in transcriptional activation by activator protein PhoB in Escherichia coli.

10.1101/gad.7.1.149

Crossref journal-article

Cold Spring Harbor Laboratory

Genes & Development (246)

Abstract

Transcription of the genes belonging to the phosphate (pho) regulon in Escherichia coli, which are induced by phosphate starvation, requires the specific activator protein PhoB in addition to the RNA polymerase holoenzyme containing the major sigma-factor sigma 70. To study the mechanism of transcriptional activation and identify the subunit of RNA polymerase involved in specific interaction with PhoB, we attempted to isolate rpoA and rpoD mutants that are specifically defective in the expression of the pho genes. We isolated two rpoD mutants with such properties, but no rpoA mutant with similar properties. The rpoD mutations altered amino acids within and near the first helix of the putative helix-turn-helix (HTH) motif in the carboxy-terminal region of sigma 70. Activities of the pho promoters in vivo were severely reduced in these mutants, whereas those of the PhoB-independent promoters were affected only marginally at most. The reconstituted mutant RNA polymerase holoenzymes were severely defective in transcribing the pstS gene, one of the pho genes, whereas they were efficient in transcribing the PhoB-independent promoters. Phosphorylated PhoB, which binds to the pho promoters with high affinity, mediated the specific binding of the wild-type holoenzyme to the pstS promoter, but it did not mediate the binding of the mutant holoenzymes. These results suggest that PhoB promotes specific interaction between RNA polymerase and the pho promoters for transcriptional activation, and the first helix of the putative HTH motif plays an essential role in the interaction, probably by making direct contact with PhoB.

Bibliography

Makino, K., Amemura, M., Kim, S. K., Nakata, A., & Shinagawa, H. (1993). Role of the sigma 70 subunit of RNA polymerase in transcriptional activation by activator protein PhoB in Escherichia coli. Genes & Development, 7(1), 149â160.

Authors 5

K Makino (first)
M Amemura (additional)
S K Kim (additional)
A Nakata (additional)
H Shinagawa (additional)

References 37 Referenced 132

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10.1128/jb.169.4.1379-1385.1987 / J. Bacteriol. / Isolation of mutations in the Î± operon of Escherichia coli that suppress the transcriptional defect conferred by a mutation in the porin regulatory gene envZ. (1987)
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10.1016/0092-8674(91)90553-B
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{'key': '2021111418065291000_7.1.149.14', 'first-page': '459', 'article-title': 'The physical map of the whole E. coli chromosome: Application of a new strategy for rapid analysis and sorting of a large genomic library.', 'volume': '50', 'year': '1987', 'journal-title': 'Cell'} / Cell / The physical map of the whole E. coli chromosome: Application of a new strategy for rapid analysis and sorting of a large genomic library. (1987)
10.1128/jb.173.23.7511-7518.1991 / J. Bacteriol. / Mutations in rpoA affect expression of anaerobically regulated genes in Salmonella typhimurium. (1991)
10.1021/bi00574a034
10.1007/BF00331115
10.1016/0022-2836(86)90073-2
10.1016/0022-2836(88)90093-9
10.1016/0022-2836(89)90131-9
10.1128/jb.173.8.2665-2672.1991 / J. Bacteriol. / Molecular analysis of the cryptic and functional phn operons for phosphonate use in Escherichia coli K-12. (1991)
10.1016/0022-2836(87)90489-X
Miller, J.H. 1972. Experiments in molecular genetics. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York.
Nakata, A., M. Amemura, K. Makino, and H. Shinagawa. 1987. Genetic and biochemical analysis of the phosphate-specific transport system in Escherichia coli. In Phosphate metabolism and cellular regulation in microorganisms (ed. A. Torriani-Gorini, F.G. Rothman, S. Silver, A. Wright, and E. Yagil), pp. 150â155. American Society for Microbiology, Washington, D.C.
{'key': '2021111418065291000_7.1.149.25', 'first-page': '912', 'article-title': 'phs locus of Escherichia coli, a mutation causing pleiotropic lesions in metabolism, is an rpoA allele.', 'volume': '164', 'year': '1985', 'journal-title': 'J. Bacteriol.'} / J. Bacteriol. / phs locus of Escherichia coli, a mutation causing pleiotropic lesions in metabolism, is an rpoA allele. (1985)
Shinagawa, H., K. Makino, M. Amemura, and A. Nakata. 1987. Structure and function of the regulatory genes for the phosphate regulon in Escherichia coli. In Phosphate metabolism and cellular regulation in microorganisms (ed. A. Torriani-Gorini, F.G. Rothman, S. Silver, A. Wright, and E. Yagil), pp. 20â25. American Society for Microbiology, Washington, D.C.
10.1016/0022-2836(88)90088-5
10.1016/0022-2836(89)90568-8
10.1128/mr.53.1.1-24.1989 / Microbiol. Rev. / A collection of strains containing genetically linked alternating antibiotic resistance elements for genetic mapping of Escherichia coli. (1989)
10.1128/jb.173.23.7501-7510.1991 / J. Bacteriol. / Suppressor mutations in rpoA suggest that OmpR controls transcription by direct interaction with the Î± subunit of RNA polymerase. (1991)
10.1128/mr.53.4.450-490.1989 / Microbiol. Rev. / Protein phosphorylation and regulation of adaptive responses in bacteria. (1989)
10.1016/0022-2836(86)90385-2
10.1073/pnas.72.7.2770
10.1111/j.1365-2958.1991.tb01980.x / Mol. Microbiol. / Escherichia coli rpoA mutation which impairs transcription of positively regulated systems. (1991)
10.1016/S0022-2836(05)80345-6
10.1128/jb.171.10.5601-5606.1989 / J. Bactriol. / Regulation of the phosphate regulon of Escherichia coli: Analysis of mutant phoB and phoR genes causing different phenotypes. (1989)
10.1016/0378-1119(85)90120-9

Dates

Type	When
Created	18 years, 2 months ago (June 5, 2007, 5:15 p.m.)
Deposited	3 years, 9 months ago (Nov. 14, 2021, 9:07 p.m.)
Indexed	3 months, 2 weeks ago (May 21, 2025, 2:55 a.m.)
Issued	32 years, 8 months ago (Jan. 1, 1993)
Published	32 years, 8 months ago (Jan. 1, 1993)
Published Online	32 years, 8 months ago (Jan. 1, 1993)
Published Print	32 years, 8 months ago (Jan. 1, 1993)

Funders 0

None

BibTeX

@article{Makino_1993, title={Role of the sigma 70 subunit of RNA polymerase in transcriptional activation by activator protein PhoB in Escherichia coli.}, volume={7}, ISSN={1549-5477}, url={http://dx.doi.org/10.1101/gad.7.1.149}, DOI={10.1101/gad.7.1.149}, number={1}, journal={Genes & Development}, publisher={Cold Spring Harbor Laboratory}, author={Makino, K and Amemura, M and Kim, S K and Nakata, A and Shinagawa, H}, year={1993}, month=jan, pages={149–160} }

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  "abstract": "<jats:p>Transcription of the genes belonging to the phosphate (pho) regulon in Escherichia coli, which are induced by phosphate starvation, requires the specific activator protein PhoB in addition to the RNA polymerase holoenzyme containing the major sigma-factor sigma 70. To study the mechanism of transcriptional activation and identify the subunit of RNA polymerase involved in specific interaction with PhoB, we attempted to isolate rpoA and rpoD mutants that are specifically defective in the expression of the pho genes. We isolated two rpoD mutants with such properties, but no rpoA mutant with similar properties. The rpoD mutations altered amino acids within and near the first helix of the putative helix-turn-helix (HTH) motif in the carboxy-terminal region of sigma 70. Activities of the pho promoters in vivo were severely reduced in these mutants, whereas those of the PhoB-independent promoters were affected only marginally at most. The reconstituted mutant RNA polymerase holoenzymes were severely defective in transcribing the pstS gene, one of the pho genes, whereas they were efficient in transcribing the PhoB-independent promoters. Phosphorylated PhoB, which binds to the pho promoters with high affinity, mediated the specific binding of the wild-type holoenzyme to the pstS promoter, but it did not mediate the binding of the mutant holoenzymes. These results suggest that PhoB promotes specific interaction between RNA polymerase and the pho promoters for transcriptional activation, and the first helix of the putative HTH motif plays an essential role in the interaction, probably by making direct contact with PhoB.</jats:p>",
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