Repression of btuB gene transcription in Escherichia coliby the GadX protein

BMC Microbiology

Research Article

Wensi S Hu¹  Kin-Fu Chak²  Po-Huang Liang³  Guang-Sheng Lei⁴  Wan-Jr Syu⁴  Shiau-Ting Hu⁵ 

[1] Department of Biotechnology and Laboratory Science in Medicine, National Yang-Ming University, 11221, Taipei, Taiwan;Institute of Biochemistry, National Yang Ming University, 11221, Taipei, Taiwan;Institute of Biological Chemistry, Academia Sinica, 11529, Taipei, Taiwan;Institute of Microbiology and Immunology, School of Life Science, National Yang-Ming University, 11221, Taipei, Taiwan;Institute of Microbiology and Immunology, School of Life Science, National Yang-Ming University, 11221, Taipei, Taiwan;Department of Education and Research, Taipei City Hospital, Taipei, Taiwan;

关键词: Cobalamin;    Proton Motive Force;    Adenosylcobalamin;    ColE7 Resistance;    Promoterless lacZ Gene;   

DOI  :  10.1186/1471-2180-11-33

 received in 2010-06-18, accepted in 2011-02-11,  发布年份 2011

来源: Springer

【 摘 要 】

BackgroundBtuB (B twelve uptake) is an outer membrane protein of Escherichia coli, it serves as a receptor for cobalamines uptake or bactericidal toxin entry. A decrease in the production of the BtuB protein would cause E. coli to become resistant to colicins. The production of BtuB has been shown to be regulated at the post-transcriptional level. The secondary structure switch of 5' untranslated region of butB and the intracellular concentration of adenosylcobalamin (Ado-Cbl) would affect the translation efficiency and RNA stability of btuB. The transcriptional regulation of btuB expression is still unclear.ResultsTo determine whether the btuB gene is also transcriptionally controlled by trans-acting factors, a genomic library was screened for clones that enable E. coli to grow in the presence of colicin E7, and a plasmid carrying gadX and gadY genes was isolated. The lacZ reporter gene assay revealed that these two genes decreased the btuB promoter activity by approximately 50%, and the production of the BtuB protein was reduced by approximately 90% in the presence of a plasmid carrying both gadX and gadY genes in E. coli as determined by Western blotting. Results of electrophoretic mobility assay and DNase I footprinting indicated that the GadX protein binds to the 5' untranslated region of the btuB gene. Since gadX and gadY genes are more highly expressed under acidic conditions, the transcriptional level of btuB in cells cultured in pH 7.4 or pH 5.5 medium was examined by quantitative real-time PCR to investigate the effect of GadX. The results showed the transcription of gadX with 1.4-fold increase but the level of btuB was reduced to 57%.ConclusionsThrough biological and biochemical analysis, we have demonstrated the GadX can directly interact with btuB promoter and affect the expression of btuB. In conclusion, this study provides the first evidence that the expression of btuB gene is transcriptionally repressed by the acid responsive genes gadX and gadY.

【 授权许可】

Unknown   
© Lei et al; licensee BioMed Central Ltd. 2011. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.