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Crucial roles of both flanking sequences in silencing of the hilA promoter in Salmonella enterica.
J Mol Biol. 2006 Mar 24; 357(2):373-86.JM

Abstract

The hilA gene on the Salmonella enterica pathogenicity island-1 encodes the key transcriptional regulator of host cell invasion. Transcription of hilA is regulated by numerous physiological signals, including repression under low osmolarity conditions. To investigate the osmotic control of hilA transcription, promoter truncations that remove sequences flanking the hilA promoter were examined. Expression of the minimal hilA core promoter (-55 to +90, relative to the transcription start site) was 57-times higher than the intact promoter (-242 to +505) in the absence of osmotic stress. Both flanking sequences contributed to the strong silencing effect, which was greatly relieved by the simultaneous loss of the two nucleoid-structuring proteins, H-NS and Hha. Mobility-shift assays revealed the presence of binding sites for the H-NS and Hha proteins, both upstream and downstream of the promoter. Either flanking region depressed expression when it was placed downstream of the lacUV5 promoter, and this inhibition was increased when the other flanking sequence was present upstream of the promoter. These results show that the hilA promoter is highly active without other transcription regulators. Its high activity is strongly depressed in low osmolarity conditions by the nucleoid-structuring proteins H-NS and Hha, possibly by formation of a repressive DNA loop. The hilA activators, HilD and HilC appear to overcome effects of downstream silencing region and disrupt repressive DNA loop. Action of activators requires contact with RNA polymerase from their DNA binding site, centered at position -77, relative to the hilA transcription start site.

Authors+Show Affiliations

Department of Microbiology, University of Virginia School of Medicine, Charlottesville, VA 22908-0734, USA. io4n@virginia.eduNo affiliation info available

Pub Type(s)

Journal Article
Research Support, N.I.H., Extramural

Language

eng

PubMed ID

16443238

Citation

Olekhnovich, Igor N., and Robert J. Kadner. "Crucial Roles of Both Flanking Sequences in Silencing of the hilA Promoter in Salmonella Enterica." Journal of Molecular Biology, vol. 357, no. 2, 2006, pp. 373-86.
Olekhnovich IN, Kadner RJ. Crucial roles of both flanking sequences in silencing of the hilA promoter in Salmonella enterica. J Mol Biol. 2006;357(2):373-86.
Olekhnovich, I. N., & Kadner, R. J. (2006). Crucial roles of both flanking sequences in silencing of the hilA promoter in Salmonella enterica. Journal of Molecular Biology, 357(2), 373-86.
Olekhnovich IN, Kadner RJ. Crucial Roles of Both Flanking Sequences in Silencing of the hilA Promoter in Salmonella Enterica. J Mol Biol. 2006 Mar 24;357(2):373-86. PubMed PMID: 16443238.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Crucial roles of both flanking sequences in silencing of the hilA promoter in Salmonella enterica. AU - Olekhnovich,Igor N, AU - Kadner,Robert J, Y1 - 2006/01/19/ PY - 2005/11/05/received PY - 2005/12/22/revised PY - 2006/01/05/accepted PY - 2006/1/31/pubmed PY - 2006/4/28/medline PY - 2006/1/31/entrez SP - 373 EP - 86 JF - Journal of molecular biology JO - J Mol Biol VL - 357 IS - 2 N2 - The hilA gene on the Salmonella enterica pathogenicity island-1 encodes the key transcriptional regulator of host cell invasion. Transcription of hilA is regulated by numerous physiological signals, including repression under low osmolarity conditions. To investigate the osmotic control of hilA transcription, promoter truncations that remove sequences flanking the hilA promoter were examined. Expression of the minimal hilA core promoter (-55 to +90, relative to the transcription start site) was 57-times higher than the intact promoter (-242 to +505) in the absence of osmotic stress. Both flanking sequences contributed to the strong silencing effect, which was greatly relieved by the simultaneous loss of the two nucleoid-structuring proteins, H-NS and Hha. Mobility-shift assays revealed the presence of binding sites for the H-NS and Hha proteins, both upstream and downstream of the promoter. Either flanking region depressed expression when it was placed downstream of the lacUV5 promoter, and this inhibition was increased when the other flanking sequence was present upstream of the promoter. These results show that the hilA promoter is highly active without other transcription regulators. Its high activity is strongly depressed in low osmolarity conditions by the nucleoid-structuring proteins H-NS and Hha, possibly by formation of a repressive DNA loop. The hilA activators, HilD and HilC appear to overcome effects of downstream silencing region and disrupt repressive DNA loop. Action of activators requires contact with RNA polymerase from their DNA binding site, centered at position -77, relative to the hilA transcription start site. SN - 0022-2836 UR - https://www.unboundmedicine.com/medline/citation/16443238/Crucial_roles_of_both_flanking_sequences_in_silencing_of_the_hilA_promoter_in_Salmonella_enterica_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0022-2836(06)00025-8 DB - PRIME DP - Unbound Medicine ER -