Investigating Transcription Factors Involved in the Coordinated Regulation of Adherence and Motility in Uropathogenic Escherichia coli
Luterbach, Courtney
2018
Abstract
Uropathogenic Escherichia coli (UPEC) are the main cause of uncomplicated urinary tract infections (UTIs), one of the most common bacterial infections in humans. Both adherence and motility are critical for productive colonization of the urinary tract. However, the mechanisms involved in coordinating the transition between adherence and motility are not well characterized. Pyelonephritis-associated pili (Pap), or P-fimbriae, bind the P-blood group antigen located on human kidney epithelial cells and erythrocytes, and UPEC strains harboring the pap operon are more likely to cause pyelonephritis. In this dissertation, a signature-tagged mutagenesis screen identified a P-fimbrial gene (papC) and 18 other genes as being among those required for full fitness of the cystitis isolate E. coli F11. Additionally, P fimbriae were confirmed by Molecular Koch’s postulates as a virulence factor for the pyelonephritis isolate E. coli CFT073 in the murine model of UTI. The production of P fimbriae is coordinated with the repression of swimming motility. Unlike the majority of other fimbrial operons, the 3’ end of the pap operon encodes a MarR-like transcription factor, PapX. Using SELEX and high-throughput sequencing, our lab has previously shown that PapX binds to a 29-bp palindromic DNA sequence located upstream of flhDC, the master regulator of flagellar gene expression, and thereby represses motility. However, the UPEC strain CFT073 carries both papX and a homolog focX, located in the foc operon encoding F1C fimbriae. In this dissertation, the dose-effects of these “X” genes on flagellar gene expression and cross-talk between focX and papX were investigated. Similar to PapX, the production of FocX was shown to repress flhDC transcription. Furthermore, the deletion of PapX and FocX in CFT073 resulted in a subtle, but not statistically significant, decrease in kidney colonization in the ascending murine model of UTI. Using 5'RACE, a proximal promoter was located upstream of both focX and papX, suggesting that these genes are transcribed independently from their fimbrial operons and therefore may be regulated by different transcription factors. Indeed, the deletion of focX resulted in increased expression of papX but had no effect on papA expression. Thus, cross-talk between "X" genes may provide a mechanism to mediate fine-tune coordinated transitions between motility and adherence. Similar to PapX and FocX, the transcription factor TosR is a critical component of the regulatory network linking adherence and motility in UPEC. TosR, encoded by the type one secretion (tos) operon, has previously been shown to function as a transcriptional repressor of the pap operon and as a dual regulator of the tos operon. The tos operon also encodes the non-fimbrial adhesin TosA, which is predominately expressed during murine UTI, binds to kidney epithelial cells, and promotes survival during invasive infections. In this dissertation, TosR was demonstrated to also regulate the expression of genes involved with adhesins, including P, F1C, and Auf; nitrate/nitrite transport; microcin secretion; and biofilm formation. Altogether, these studies provide an in-depth characterization of three transcription factors (PapX, FocX, and TosR) and their contribution to the coordinated regulation of motility and adherence in UPEC.Subjects
Adherence Motility UPEC Gene Regulation
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