| 1. |
- Greene, Sarah E., et al.
(author)
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Pilicide ec240 Disrupts Virulence Circuits in Uropathogenic Escherichia coli
- 2014
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In: mBio. - 2161-2129 .- 2150-7511. ; 5:6, s. UNSP e02038-
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Journal article (peer-reviewed)abstract
- Chaperone-usher pathway (CUP) pili are extracellular organelles produced by Gram-negative bacteria that mediate bacterial pathogenesis. Small-molecule inhibitors of CUP pili, termed pilicides, were rationally designed and shown to inhibit type 1 or P piliation. Here, we show that pilicide ec240 decreased the levels of type 1, P, and S piliation. Transcriptomic and proteomic analyses using the cystitis isolate UTI89 revealed that ec240 dysregulated CUP pili and decreased motility. Paradoxically, the transcript levels of P and S pilus genes were increased during growth in ec240, even though the level of P and S piliation decreased. In contrast, the most downregulated transcripts after growth in ec240 were from the type 1 pilus genes. Type 1 pilus expression is controlled by inversion of the fimS promoter element, which can oscillate between phase on and phase off orientations. ec240 induced the fimS phase off orientation, and this effect was necessary for the majority of ec240's inhibition of type 1 piliation. ec240 increased levels of the transcriptional regulators SfaB and PapB, which were shown to induce the fimS promoter phase off orientation. Furthermore, the effect of ec240 on motility was abolished in the absence of the SfaB, PapB, SfaX, and PapX regulators. In contrast to the effects of ec240, deletion of the type 1 pilus operon led to increased S and P piliation and motility. Thus, ec240 dysregulated several uropathogenic Escherichia coli (UPEC) virulence factors through different mechanisms and independent of its effects on type 1 pilus biogenesis and may have potential as an antivirulence compound. IMPORTANCE CUP pili and flagella play active roles in the pathogenesis of a variety of Gram-negative bacterial infections, including urinary tract infections mediated by UPEC. These are extremely common infections that are often recurrent and increasingly caused by antibiotic-resistant organisms. Preventing piliation and motility through altered regulation and assembly of these important virulence factors could aid in the development of novel therapeutics. This study increases our understanding of the regulation of these virulence factors, providing new avenues by which to target their expression.
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| 2. |
- Spaulding, Caitlin N., et al.
(author)
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Functional role of the type 1 pilus rod structure in mediating host-pathogen interactions
- 2018
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In: eLIFE. - : eLife Sciences Publications. - 2050-084X. ; 7
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Journal article (peer-reviewed)abstract
- Uropathogenic E. coli (UPEC), which cause urinary tract infections (UTI), utilize type 1 pili, a chaperone usher pathway (CUP) pilus, to cause UTI and colonize the gut. The pilus rod, comprised of repeating FimA subunits, provides a structural scaffold for displaying the tip adhesin, FimH. We solved the 4.2 Å resolution structure of the type 1 pilus rod using cryo-electron microscopy. Residues forming the interactive surfaces that determine the mechanical properties of the rod were maintained by selection based on a global alignment of fimA sequences. We identified mutations that did not alter pilus production in vitro but reduced the force required to unwind the rod. UPEC expressing these mutant pili were significantly attenuated in bladder infection and intestinal colonization in mice. This study elucidates an unappreciated functional role for the molecular spring-like property of type 1 pilus rods in host-pathogen interactions and carries important implications for other pilus-mediated diseases.
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| 3. |
- Zheng, Weili, et al.
(author)
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Cryo-Em Structure of Type 1 Pilus
- 2018
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In: Biophysical Journal. - : Biophysical Society. - 0006-3495 .- 1542-0086. ; 114:3, s. 370a-370a
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Journal article (other academic/artistic)abstract
- Urinary tract infections (UTIs) are caused by a wide range of pathogens, but the most common causative agent of UTIs is uropathogenic Escherichia coli (UPEC). Virtually all uropathogenic strains of E. coli encode filamentous surface adhesive organelles called type 1 pili, which are a subset of Chaperone-usher pathway (CUP) pili. CUP pili are also ubiquitously expressed on the surface of many other Gram-negative bacterial pathogens. They are important virulence factors facilitating host-pathogen interactions that are crucial for the establishment and persistence of an infection, and involved in regulating other key processes such as biofilm formation. We have solved the 4.2 Å resolution cryo-EM structure of the type 1 pilus, which was present as a background contaminant in a prep of type 4 pili. We have taken advantage of the strength of cryo-EM to separate different molecules and conformations present in solution to show that filament images which might otherwise have been discarded as a contaminant can actually be used to build an atomic model. The model reveals the residues that allow a long chain of FimA subunits, linked by the insertion of a β-strand of one subunit into the β-sheet of an adjacent subunit, to coil into a rigid rod. We show that site-specific mutation of these residues reduces the force needed to unwind the rod. Strikingly, one mutation (A22R) which showed the greatest reduction in unwinding force, eliminated bladder infections in a mouse model. This is presumably due to the fact that the altered mechanics of the A22R pilus rod cannot withstand the shear forces due to urinary flow in the bladder and bacteria harboring this mutation are cleared from the bladder. This provides new insights into the important role of pili mechanics in bacterial pathogenesis.
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