Role of phosphate-specific transport system in the expression of LEE genes of Enterohemorrhagic Escherichia coli (EHEC)

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Role of phosphate-specific transport system in the expression of LEE genes of Enterohemorrhagic Escherichia coli (EHEC)

 

Author: Ho, Wing-tak Sharon
Title: Role of phosphate-specific transport system in the expression of LEE genes of Enterohemorrhagic Escherichia coli (EHEC)
Degree: M.Sc.
Year: 2010
Subject: Hong Kong Polytechnic University -- Dissertations
Escherichia coli O157:H7 -- Genetic aspects
Phosphorus -- Metabolism.
Biological transport.
Department: Dept. of Health Technology and Informatics
Pages: viii, 45 leaves : col. ill. ; 30 cm.
InnoPac Record: http://library.polyu.edu.hk/record=b2352682
URI: http://theses.lib.polyu.edu.hk/handle/200/5650
Abstract: Enterohemorrhagic Escherichia coli (EHEC) is an emerging zoonotic pathogen associated with human diseases ranging from mild diarrhea to hemolytic uremic syndrome. Virulence factor of EHEC is intimin, an important bacterial adhesin, which is associated with intimate attachment of EHEC to host cells. The intimin is encoded by the eaeA gene, which is located in the locus of enterocyte effacement (LEE) island. Intimin will activate a series of signal transduction and causes accumulation of actin at the attachment site, thus resulting in the formation of attaching/effacing lesion (A/E) in the host intestinal surface. The LEE island consists of LEE1-5. LEE1-3 encode proteins involved in the biogenesis of a type III secretion system (TTSS), a complex multiprotein organelles, which permits translocation of effector proteins from the bacterial cell to envelope into the eukaryotic cells cytosol. LEE4 encodes the Esp protein, for example espA which plays an important function for mammalian cell infection: it acts as sole constituent of hollow filament for translocator protein EspA. In this project, an EHEC strain with a disrupted pstC was studied for the changes in LEE gene expression. Expression of LEE genes and LEE regulators were compared using quantitative real-time polymerase chain reaction (RT-PCR) and then by calculation of ΔΔCT value to detect if there were any LEE genes or LEE regulators expression changes. Investigation of genes in this project included LEE-encoded regulators: ler (LEE1), grlA (between LEE1 & 2), grlR (between LEE1 & 2). Non-LEE-encoded regulators: qseA, sdiA, hns, pchA, pchB. LEE genes: escJ (LEE2), escV (LEE3), espA (LEE4), eaeA (LEE5), tir (LEE5). rpoA was employed as a reference gene. When compared to the EHEC wild-type strain, the pstC mutant showed a reduction in gene expression of four LEE regulators namely, ler, grlR, grlA and pchA (ΔΔCT ranged from -5.49 to -1.96). Other LEE genes being down-regulated included escV, espA and eaeA. (ΔΔCT ranged from -7.19 to -5.01). In this study, it was shown that gene disruption in pstC caused a reduction of gene expression of LEE regulators and effectors. Further studies on the effect of pstC disruption of EHEC adherence properties are needed.

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