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Science 10 November 1995:
Vol. 270. no. 5238, pp. 992 - 994
DOI: 10.1126/science.270.5238.992
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Reports

Converting Escherichia coli RNA Polymerase into an Enhancer-Responsive Enzyme: Role of an NH2-Terminal Leucine Patch in 54

Jonathan T. Wang,  Adeela Syed,  Mingli Hsieh (1),  Jay D. Gralla (2)

The protein 54 associates with Escherichia coli core RNA polymerase to form a holoenzyme that binds promoters but is inactive in the absence of enhancer activation. Here, mutants of 54 enabled polymerases to transcribe without enhancer protein and adenosine triphosphate. The mutations are in leucines within the NH2-terminal glutamine-rich domain of 54. Multiple leucine substitutions mimicked the effect of enhancer protein, which suggests that the enhancer protein functions to disrupt a leucine patch. The results indicate that 54 acts both as an inhibitor of polymerase activity and as a receptor that interacts with enhancer protein to overcome this inhibition, and that these two activities jointly confer enhancer responsiveness.


Department of Chemistry and Biochemistry and Molecular Biology Institute, University of California, Los Angeles, CA 90095, USA.
(1) Present address: Institute of Medicine, Chung Shan Medical and Dental College, Taichung, Taiwan.
(2) To whom correspondence should be addressed.


THIS ARTICLE HAS BEEN CITED BY OTHER ARTICLES:
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A Role for the Conserved GAFTGA Motif of AAA+ Transcription Activators in Sensing Promoter DNA Conformation.
A. E. Dago, S. R. Wigneshweraraj, M. Buck, and E. Morett (2007)
J. Biol. Chem. 282, 1087-1097
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Interactions of regulated and deregulated forms of the {sigma}54 holoenzyme with heteroduplex promoter DNA.
W. Cannon, S. R. Wigneshweraraj, and M. Buck (2002)
Nucleic Acids Res. 30, 886-893
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Integration of Global Regulation of Two Aromatic-Responsive {sigma}54-Dependent Systems: a Common Phenotype by Different Mechanisms.
C. C. Sze, L. M. D. Bernardo, and V. Shingler (2002)
J. Bacteriol. 184, 760-770
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Action of prokaryotic enhancer over a distance does not require continued presence of promoter-bound {sigma}54 subunit.
V. Bondarenko, Y. Liu, A. Ninfa, and V. M. Studitsky (2002)
Nucleic Acids Res. 30, 636-642
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DNA supercoiling allows enhancer action over a large distance.
Y. Liu, V. Bondarenko, A. Ninfa, and V. M. Studitsky (2001)
PNAS
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Binding of transcriptional activators to sigma 54 in the presence of the transition state analog ADP-aluminum fluoride: insights into activator mechanochemical action.
M. Chaney, R. Grande, S. R. Wigneshweraraj, W. Cannon, P. Casaz, M.-T. Gallegos, J. Schumacher, S. Jones, S. Elderkin, A. E. Dago, et al. (2001)
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In vitro roles of invariant helix-turn-helix motif residue R383 in {{sigma}}54 ({{sigma}}N).
S. R. Wigneshweraraj, A. Ishihama, and M. Buck (2001)
Nucleic Acids Res. 29, 1163-1174
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Transcription Initiation-Defective Forms of sigma 54 That Differ in Ability To Function with a Heteroduplex DNA Template.
M. T. Kelly, J. A. Ferguson III, and T. R. Hoover (2000)
J. Bacteriol. 182, 6503-6508
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The role of Region II in the RNA polymerase {sigma} factor {sigma}N ({sigma}54).
E. Southern and M. Merrick (2000)
Nucleic Acids Res. 28, 2563-2570
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Low Resolution Structure of the sigma 54 Transcription Factor Revealed by X-ray Solution Scattering.
D. I. Svergun, M. Malfois, M. H. J. Koch, S. R. Wigneshweraraj, and M. Buck (2000)
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Role of Tissue Plasminogen Activator Receptor LRP in Hippocampal Long-Term Potentiation.
M. Zhuo, D. M. Holtzman, Y. Li, H. Osaka, J. DeMaro, M. Jacquin, and G. Bu (2000)
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Regulation of Sigma 54-Dependent Transcription by Core Promoter Sequences: Role of -12 Region Nucleotides.
L. Wang, Y. Guo, and J. D. Gralla (1999)
J. Bacteriol. 181, 7558-7565
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The hydrophobic heptad repeat in Region III of Escherichia coli transcription factor sigma 54 is essential for core RNA polymerase binding.
M. Hsieh, H.-M. Hsu, S.-F. Hwang, F.-C. Wen, J.-S. Yu, C.-C. Wen, and C. Li (1999)
Microbiology 145, 3081-3088
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Functions of the sigma 54 Region I in Trans and Implications for Transcription Activation.
M.-T. Gallegos, W. V. Cannon, and M. Buck (1999)
J. Biol. Chem. 274, 25285-25290
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Amino-terminal sequences of sigma N (sigma 54) inhibit RNA polymerase isomerization.
W. Cannon, M.-T. Gallegos, P. Casaz, and M. Buck (1999)
Genes & Dev. 13, 357-370
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Identification of an N-Terminal Region of Sigma 54 Required for Enhancer Responsiveness.
A. Syed and J. D. Gralla (1998)
J. Bacteriol. 180, 5619-5625
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Multiple In Vivo Roles for the -12-Region Elements of Sigma 54 Promoters.
L. Wang and J. D. Gralla (1998)
J. Bacteriol. 180, 5626-5631
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Y. Guo and J. D. Gralla (1998)
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A bacterial ATP-dependent, enhancer binding protein that activates the housekeeping RNA polymerase.
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A. KUMAR, A. GROVE, G.A. KASSAVETIS, and E.P. GEIDUSCHEK (1998)
Cold Spring Harb Symp Quant Biol 63, 121-131
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The Bacterial Enhancer-binding Protein NtrC as a Molecular Machine.
I. ROMBEL, A. NORTH, I. HWANG, C. WYMAN, and S. KUSTU (1998)
Cold Spring Harb Symp Quant Biol 63, 157-166
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Probing the assembly of transcription initiation complexes through changes in sigma N protease sensitivity.
P. Casaz and M. Buck (1997)
PNAS 94, 12145-12150
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Multiple pathways to bypass the enhancer requirement of sigma 54 RNA polymerase: Roles for DNA and protein determinants.
J. T. Wang, A. Syed, and J. D. Gralla (1997)
PNAS 94, 9538-9543
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Two domains within sigma N (sigma 54) cooperate for DNA binding.
W. V. Cannon, M. K. Chaney, X.-Y. Wang, and M. Buck (1997)
PNAS 94, 5006-5011
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The Transcription Initiation Pathway of Sigma 54Mutants That Bypass the Enhancer Protein Requirement. IMPLICATIONS FOR THE MECHANISM OF ACTIVATION.
J. T. Wang and J. D. Gralla (1996)
J. Biol. Chem. 271, 32707-32713
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Genetic Evidence for Interdomain Regulation of the Phenol-responsive &sfgr;54-dependent Activator DmpR.
L. Ng, E. O'Neill, and V. Shingler (1996)
J. Biol. Chem. 271, 17281-17286
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Sequences within the DNA Cross-linking Patch of sigma 54 Involved in Promoter Recognition, sigma Isomerization, and Open Complex Formation.
M. Chaney, M. Pitt, and M. Buck (2000)
J. Biol. Chem. 275, 22104-22113
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DNA Melting within a Binary sigma 54-Promoter DNA Complex.
W. Cannon, M.-T. Gallegos, and M. Buck (2001)
J. Biol. Chem. 276, 386-394
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Roles for the C-terminal Region of Sigma 54 in Transcriptional Silencing and DNA Binding.
L. Wang and J. D. Gralla (2001)
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A Gal4-sigma 54 Hybrid Protein That Functions as a Potent Activator of RNA Polymerase II Transcription in Yeast.
B.-S. Chen, Z.-W. Sun, and M. Hampsey (2001)
J. Biol. Chem. 276, 23881-23887
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DNA supercoiling allows enhancer action over a large distance.
Y. Liu, V. Bondarenko, A. Ninfa, and V. M. Studitsky (2001)
PNAS 98, 14883-14888
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