Functional Genomic Studies of Uropathogenic Escherichia coli and Host Urothelial Cells when Intracellular Bacterial Communities Are Assembled

Abstract

Uropathogenic Escherichia coli (UPEC), the principal cause of urinary tract infection in women, colonizes the gut as well as the genitourinary tract. Studies of mice inoculated with UTI89, a sequenced isolate, have revealed a complex life cycle that includes formation of intracellular bacterial communities (IBCs) in bladder urothelial cells. To understand how UPEC adapts to life in IBCs, we have used GeneChips and/or quantitative reverse transcriptase PCR to study UTI89 recovered from the distal gut of gnotobiotic mice and from IBCs harvested by laser capture microdissection from the bladder urothelium of infected C3H/HeJ female mice. Host responses were characterized in laser capture microdissected urothelial cells that do or do not contain IBCs. The results reveal components of ferric iron acquisition systems in UTI89 that are expressed at significantly higher levels in IBCs compared with the intestine, including the hemin receptor chuA (1,390 ؎ 188-fold). Localized urothelial responses to IBCs help oppose bacterial salvage of host cell iron (e.g. up-regulation of Tfrc (transferrin receptor) and Lcn2 (lipocalin 2)), facilitate glucose import (e.g. Hk2 (hexokinase 2)), and maintain epithelial structural integrity (e.g. Ivl (involucrin) and Sbsn (suprabasin)). ⌬chuA mutants produce significantly smaller IBCs compared with wild type UTI89. This difference was not observed in strains lacking sitA (ABC-type iron/manganese transporter subunit), iroN (salmochelin receptor), hlyA (␣-hemolysin), or entF (enterobactin synthetase subunit). Together, these studies indicate that heme-and siderophoreassociated iron play key roles in IBC development and provide a series of microbial and host biomarkers for comparing UPEC strains isolated from humans.

Innate Transcriptional Networks Activated in Bladder in Response to Uropathogenic Escherichia coli Drive Diverse Biological Pathways and Rapid Synthesis of IL-10 for Defense against Bacterial Urinary Tract Infection

“…Our approach has uncovered a broad range of host factors not previously known to regulate innate defense to bacterial UTI. Several genes identified in this study are consistent with prior reports that profiled mouse bladder previously and mapped responses in foci of uroepithelium infected with intracellular UPEC pods, which displayed localized responses (50,57). The whole tissue approach we have used gives broader definition to the total bladder transcriptome of UTI.…”

“…Notably, many of the responses identified in the whole bladder transcriptome presented in this work are consistent with those observed in localized responses to intracellular bacterial communities in bladder urothelial cells, as previously reported (57). Of particular note were localized urothelial responses to intracellular bacterial communities such as those directed at opposing UPEC salvage of host cell iron and facilitation of glucose import and epithelial structural integrity (57). Many of these responses are consistent with the genes, such as lipocalin 2 (Lcn2) and hexokinase 2 (Hk2), identified as highly upregulated in the whole bladder transcriptome of UPEC cystitis in this study.…”

What does it take to stick around?

“…32,33 In UPEC, a metabolomic study by Henderson et al demonstrated a prevalence of yersiniabactin in UPEC vs. coincident rectal isolates, indicating a role for this siderophore during pathogenesis. 31 Consistent with this hypothesis, yersiniabactin biosynthesis genes have been shown to be highly expressed in IBCs in a murine model of infection 36 (Hadjifrangiskou et al, unpublished). More recent studies have demonstrated that disruption of salicylate production by the yersiniabactin biosynthesis pathway in UPEC results in dramatic loss of UPEC pellicle biofilm, which is restored upon exogenous addition of micromolar concentrations of salicylate (Henderson and Hung et al, unpublished data).…”

Iron induces bimodal population development by Escherichia coli

“…Growth in urine induces expression of a variety of iron acquisition systems in UPEC, demonstrating that urine is an iron-limiting environment (47). Additionally, IBC formation correlates with induction of genes involved in acquiring iron from heme and siderophores (48). Because iron and biofilm formation both play a significant role in UPEC pathogenesis, it is tempting to speculate that the iron-induced biofilm formation we describe here may contribute to UPEC virulence.…”