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Functional Analysis of LysR Family vc2103 and Its Target Gene in V.choleare

Author: JinJing
Tutor: ZhuJun
School: Nanjing Agricultural College
Course: Microbiology
Keywords: Vibrio cholera LysR-type transcriptional regulator vc2103 rifampin murein hydrolase
CLC: R346
Type: Master's thesis
Year: 2011
Downloads: 4
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Abstract


Cholera is an old and prevalent sweepingly acute intestinal diseases caused by the gram-negative bacterial Vibrio cholerae. Extensive studies have shown that the ability of V. cholerae strains to cause severe enteric infection in humans dependents on the expression of cholera toxin and a pilus colonization factor known as the toxin-coregulated pilus.LysR-type transcriptional regulators are thoroughly studied as transcriptional regulators, they are highly conserved and ubiquitous amongst bacteria, constitute the largest known family of prokaryotic DNA-binding proteins, regulating a diverse set of target genes, including metabolism, cell division, quorum sensing, virulence, motility, nitrogen fixation, oxidative stress responses, toxin production, attachment, secretion and so on. Most of the LysR genes use divergent promoter to activate expression of target genes, repress their own expression. We found that about 40 LysR type of protein in Vibrio cholera by bioinformatics. In this study. Vibrio cholerae LysR family genes vc2103 was used.First, we construct Luminescence detection of strains, the study shows:LysR family gene vc2103 not only inhibits its own expression but also activate downstream gene vc2101 (effector of murein hydrolase) expression in the presence of acetic acid.In this experiment, we construct vc2103 and vc2100-2101 deletion strains through in-frame deletion. We found that gene vc2103 and vc2100-2101 deletion does not affect the normal growth of Vibrio cholera. Howerve, in the presence of acetic acid, wild-type strains and vc2103 mutant declined faster on the late growth phase. The results showed that virulence gene expression and colonization ability was not affected in vc2103 mutant through Western blot and intestinal colonization in infant mice experiments, it indicates gene vc2103 and virulence of Vibrio cholera does not matter. In the presence of acetic acid, the wild-type strains and vc2103 mutant enhanced the sensitivity of rifampin compared with non-acetic acid.When vc2103 downstream gene vc2100 and vc2101 were deleted, the mutant significantly decreased the sensitivity of rifampin compared with wild type strain. In the presence of acetic acid, the wild-type strains enhanced the sensitivity of rifampin compared with non-acetic acid, but vc2100 and vc2101 deletion mutant is not much difference in the sensitivity of rifampin. This study shows vc2100 and vc2101 would enhanced the sensitivity of rifampin. Extracellular murein hydrolase expression assays have shown that vc2100 and vc2101 deletion would affect extracellular murein hydrolase normal expression.

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