Ler and H-NS, regulators controlling expression of the long polar fimbriae of Escherichia coli O157

H7

Alfredo Torres, Guillermo N. López-Sánchez, Lorena Milflores-Flores, Shilpa D. Patel, Maricarmen Rojas-López, Claudia F. Martínez De La Peña, Margarita M P Arenas-Hernández, Ygnacio Martínez-Laguna

Research output: Contribution to journalArticle

45 Citations (Scopus)

Abstract

Enterohemorrhagic Escherichia coli (EHEC) serotype O157:H7 colonizes the human intestine and is responsible for diarrheal outbreaks worldwide. Previously we showed that EHEC produces long polar fimbriae (LPF) and that maximum expression is observed during the exponential phase of growth at 37°C and pH 6.5. In this study, we analyzed the roles of several regulators in the expression of LPF using the β-galactosidase reporter system, and we found that H-NS functions as a transcriptional silencer while Ler functions as an antisilencer of LPF expression. Interestingly, deletion of the hns and ler genes in EHEC caused constitutive expression of the fusion reporter protein. Semiquantitative reverse transcription (RT)-PCR was also used to analyze LPF expression in the EHEC ler or hns mutant strain. The hns mutant exhibited an increase in lpf mRNA expression, while expression in the ler mutant was decreased, compared to that in the wild-type strain. Using primer extension analysis, we identified two potential transcriptional start sites within the regulatory region of lpf and located consensus hexamers of -10 (CAAGAT) and -35 (TTCAAA), which are commonly found in σ70-dependent promoters. Further, we determined whether H-NS and Ler interact directly with the lpf promoter region by using purified His-tagged proteins and electrophoretic mobility shift assays. Our data are the first to show direct binding interactions between the H-NS and Ler proteins within the regulatory sequence of the lpf operon. Based on the electrophoretic mobility shift assay, RT-PCR, primer extension, and β-galactosidase assay results, we concluded that the E. coli O157:H7 lpf operon possesses a promoter dependent on σ70, that H-NS binds to the regulatory sequence of lpf A and "silences" the transcription of lpf, and that Ler binds to the regulatory sequence and inhibits the action of the H-NS protein.

Original languageEnglish (US)
Pages (from-to)5916-5928
Number of pages13
JournalJournal of Bacteriology
Volume189
Issue number16
DOIs
StatePublished - Aug 2007

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Enterohemorrhagic Escherichia coli
Escherichia coli O157
Galactosidases
Electrophoretic Mobility Shift Assay
Operon
Reverse Transcription
Proteins
Polymerase Chain Reaction
Nucleic Acid Regulatory Sequences
Genetic Promoter Regions
Intestines
Disease Outbreaks
Messenger RNA
Growth
Genes

ASJC Scopus subject areas

  • Applied Microbiology and Biotechnology
  • Immunology

Cite this

Torres, A., López-Sánchez, G. N., Milflores-Flores, L., Patel, S. D., Rojas-López, M., Martínez De La Peña, C. F., ... Martínez-Laguna, Y. (2007). Ler and H-NS, regulators controlling expression of the long polar fimbriae of Escherichia coli O157: H7. Journal of Bacteriology, 189(16), 5916-5928. https://doi.org/10.1128/JB.00245-07

Ler and H-NS, regulators controlling expression of the long polar fimbriae of Escherichia coli O157 : H7. / Torres, Alfredo; López-Sánchez, Guillermo N.; Milflores-Flores, Lorena; Patel, Shilpa D.; Rojas-López, Maricarmen; Martínez De La Peña, Claudia F.; Arenas-Hernández, Margarita M P; Martínez-Laguna, Ygnacio.

In: Journal of Bacteriology, Vol. 189, No. 16, 08.2007, p. 5916-5928.

Research output: Contribution to journalArticle

Torres, A, López-Sánchez, GN, Milflores-Flores, L, Patel, SD, Rojas-López, M, Martínez De La Peña, CF, Arenas-Hernández, MMP & Martínez-Laguna, Y 2007, 'Ler and H-NS, regulators controlling expression of the long polar fimbriae of Escherichia coli O157: H7', Journal of Bacteriology, vol. 189, no. 16, pp. 5916-5928. https://doi.org/10.1128/JB.00245-07
Torres A, López-Sánchez GN, Milflores-Flores L, Patel SD, Rojas-López M, Martínez De La Peña CF et al. Ler and H-NS, regulators controlling expression of the long polar fimbriae of Escherichia coli O157: H7. Journal of Bacteriology. 2007 Aug;189(16):5916-5928. https://doi.org/10.1128/JB.00245-07
Torres, Alfredo ; López-Sánchez, Guillermo N. ; Milflores-Flores, Lorena ; Patel, Shilpa D. ; Rojas-López, Maricarmen ; Martínez De La Peña, Claudia F. ; Arenas-Hernández, Margarita M P ; Martínez-Laguna, Ygnacio. / Ler and H-NS, regulators controlling expression of the long polar fimbriae of Escherichia coli O157 : H7. In: Journal of Bacteriology. 2007 ; Vol. 189, No. 16. pp. 5916-5928.
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AU - Milflores-Flores, Lorena

AU - Patel, Shilpa D.

AU - Rojas-López, Maricarmen

AU - Martínez De La Peña, Claudia F.

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AU - Martínez-Laguna, Ygnacio

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