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    Studies on the regulation of virulence gene expression of enteropathogenic Escherichia coli

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    Author
    Gomez, Oscar Gilberto
    Advisor
    Kaper, James B.
    Date
    1993
    Type
    dissertation
    
    Metadata
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    Abstract
    Enteropathogenic Escherichia coli (EPEC) are a well established cause of infant diarrhea around the world, being reported primarily in developing countries and only sporadically in industrialized nations. Several genes associated with virulence have been identified. Chromosomal genes are associated with the ability of EPEC to intimately adhere to epithelial cells and to modify the cytoskeleton of enterocytes which results in effacement of microvilli. A gene known as eaeA is involved in intimate adherence and encodes a 94 kDa outer membrane protein (OMP) called intimin. Other genes are located in plasmids and are associated with the ability of EPEC to adhere to epithelial cells in a localized adherence pattern. Using recombinant DNA technology a region has been isolated from the EPEC plasmid which regulates the expression of eaeA. This DNA region encodes a protein of 26 kDa which has been designated plasmid-encoded regulator A (PerA). DNA sequence analysis indicates that PerA has homology with the AraC family of transcriptional regulators. Two helix-turn-helix motifs have been identified at the C-terminus of PerA, suggesting that PerA is a DNA binding protein. Expression of eaeA was tested under different environmental conditions by measuring the alkaline phosphatase activity of a eaeA::TnphoA transposon mutant of EPEC carrying or lacking the perA gene. Temperature and exponential growth favored eaeA expression in a PerA dependent fashion whereas iron proficient conditions favored eaeA expression independently of PerA. Northern blot analysis shows that PerA is a positive regulator of transcription of not only eaeA, but also of eaeB and bfpA. eaeB is a chromosomal gene that encodes a 39 kDa protein which is essential for intimate adherence. bfpA is a plasmid gene that encodes the 19.1 kDa protein structural subunit of the bundle forming pilus (BFP). BFP is a type IV fimbriae which is associated with localized adherence of EPEC to epithelial cells. In addition to eaeA, eaeB, and bfpA genes PerA may also regulate other genes as indicated by OMP analysis. Two OMPs of 50 kDa and 33 kDa are overexpressed in {dollar}perA\sp+{dollar} strains whereas the expression of a 20 kDa OMP is decreased. Differential expression of OMPs indicates that PerA may act as a positive, as well as a negative regulator of EPEC genes. DNA sequence analysis has identified two direct repeats in the promoter regions of both eaeA and eaeB genes which may serve as potential binding sites for PerA. The ability of PerA to affect the expression of a variety of virulence associated genes suggests that PerA is a global regulator of virulence gene expression which may be essential in EPEC pathogenesis.
    Description
    University of Maryland, Baltimore. Ph.D. 1993
    Keyword
    Biology, Genetics
    Biology, Microbiology
    Enteropathogenic Escherichia coli--genetics
    Enteropathogenic Escherichia coli--pathogenicity
    Gene Expression
    Identifier to cite or link to this item
    http://hdl.handle.net/10713/1623
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    Theses and Dissertations All Schools
    Theses and Dissertations School of Medicine

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