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Role of upstream activation sequences and integration host factor in transcriptional activation by the constitutively active prokaryotic enhancer-binding protein PspF.
J Mol Biol. 1997 Oct 24; 273(2):377-88.JM

Abstract

PspF, the transcriptional activator of the pspA operon of Escherichia coli, which belongs to the enhancer binding protein (EBP) family of sigma54 activator proteins, is constitutively active in an in vitro transcription assay. PspF protein, together with RNA polymerase holoenzyme containing sigma54, is required for in vitro transcription from the pspA promoter. EBP proteins are typically subject to regulation either by post-translational modification or interaction of a specific ligand with an N-terminal regulatory domain. However, unlike other members of the EBP family, PspF lacks this domain. pspA is positively regulated by IHF in vitro, and this regulation is dependent on the topology of the DNA; a linear template is much more dependent on IHF than a supercoiled template. EBP binding to upstream activating sequences (UAS) in their target promoters is mediated by the C-terminal domain which contains a helix-turn-helix DNA-binding motif. A mutant PspF protein lacking the C-terminal DNA-binding domain is active in vitro, although at much higher concentrations than the wild-type protein. In vitro transcription from pspA templates missing one or both of the UAS sites is reduced relative to wild-type templates, but is still appreciable; however, IHF acts as a negative regulator of pspA transcription on these mutant templates. Thus, PspF bound to non-specific sequences upstream of the pspA promoter can activate pspA transcription, but this activation is inhibited by IHF. These data, taken together, support the model that a precise promoter geometry is necessary for IHF to positively regulate transcription and that IHF may act to prevent activation from inappropriately spaced upstream sites.

Authors+Show Affiliations

Rockefeller University, 1230 York Avenue, New York, NY 10021, USA.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

9344746

Citation

Dworkin, J, et al. "Role of Upstream Activation Sequences and Integration Host Factor in Transcriptional Activation By the Constitutively Active Prokaryotic Enhancer-binding Protein PspF." Journal of Molecular Biology, vol. 273, no. 2, 1997, pp. 377-88.
Dworkin J, Jovanovic G, Model P. Role of upstream activation sequences and integration host factor in transcriptional activation by the constitutively active prokaryotic enhancer-binding protein PspF. J Mol Biol. 1997;273(2):377-88.
Dworkin, J., Jovanovic, G., & Model, P. (1997). Role of upstream activation sequences and integration host factor in transcriptional activation by the constitutively active prokaryotic enhancer-binding protein PspF. Journal of Molecular Biology, 273(2), 377-88.
Dworkin J, Jovanovic G, Model P. Role of Upstream Activation Sequences and Integration Host Factor in Transcriptional Activation By the Constitutively Active Prokaryotic Enhancer-binding Protein PspF. J Mol Biol. 1997 Oct 24;273(2):377-88. PubMed PMID: 9344746.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Role of upstream activation sequences and integration host factor in transcriptional activation by the constitutively active prokaryotic enhancer-binding protein PspF. AU - Dworkin,J, AU - Jovanovic,G, AU - Model,P, PY - 1997/11/5/pubmed PY - 1997/11/5/medline PY - 1997/11/5/entrez SP - 377 EP - 88 JF - Journal of molecular biology JO - J Mol Biol VL - 273 IS - 2 N2 - PspF, the transcriptional activator of the pspA operon of Escherichia coli, which belongs to the enhancer binding protein (EBP) family of sigma54 activator proteins, is constitutively active in an in vitro transcription assay. PspF protein, together with RNA polymerase holoenzyme containing sigma54, is required for in vitro transcription from the pspA promoter. EBP proteins are typically subject to regulation either by post-translational modification or interaction of a specific ligand with an N-terminal regulatory domain. However, unlike other members of the EBP family, PspF lacks this domain. pspA is positively regulated by IHF in vitro, and this regulation is dependent on the topology of the DNA; a linear template is much more dependent on IHF than a supercoiled template. EBP binding to upstream activating sequences (UAS) in their target promoters is mediated by the C-terminal domain which contains a helix-turn-helix DNA-binding motif. A mutant PspF protein lacking the C-terminal DNA-binding domain is active in vitro, although at much higher concentrations than the wild-type protein. In vitro transcription from pspA templates missing one or both of the UAS sites is reduced relative to wild-type templates, but is still appreciable; however, IHF acts as a negative regulator of pspA transcription on these mutant templates. Thus, PspF bound to non-specific sequences upstream of the pspA promoter can activate pspA transcription, but this activation is inhibited by IHF. These data, taken together, support the model that a precise promoter geometry is necessary for IHF to positively regulate transcription and that IHF may act to prevent activation from inappropriately spaced upstream sites. SN - 0022-2836 UR - https://www.unboundmedicine.com/medline/citation/9344746/Role_of_upstream_activation_sequences_and_integration_host_factor_in_transcriptional_activation_by_the_constitutively_active_prokaryotic_enhancer_binding_protein_PspF_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0022-2836(97)91317-6 DB - PRIME DP - Unbound Medicine ER -