title: Uropathogenic Escherichia coli induces extrinsic and intrinsic cascades to initiate urothelial apoptosis.
abstract: A murine model of urinary tract infection identified urothelial apoptosis as a key event in the pathogenesis mediated by uropathogenic Escherichia coli (UPEC), yet the mechanism of this important host response is not well characterized. We employed a culture model of UPEC-urothelium interactions to examine the biochemical events associated with urothelial apoptosis induced by the UPEC strain NU14. NU14 induced DNA cleavage within 5 h that was inhibited by the broad caspase inhibitor ZVAD, and urothelial caspase 3 activity was induced within 3 h of exposure to type 1 piliated NU14 and was dependent upon interactions mediated by the type 1 pilus adhesin FimH. Flow cytometry experiments using chloromethyl-X-rosamine and Indo-1 revealed FimH-dependent mitochondrial membrane depolarization and elevated [Ca(2+)](in), respectively, indicating activation of the intrinsic apoptotic pathway. Consistent with this possibility, overexpression of Bcl(XL) inhibited NU14 activation of caspase 3. Immunoblotting, caspase inhibitors, and caspase activity assays implicated both caspase 2 and caspase 8 in apoptosis, suggesting the involvement of the intrinsic and extrinsic apoptotic cascades. To reconcile the apparent activation of both extrinsic and intrinsic pathways, we examined Bid-green fluorescent protein localization and observed translocation from the cytosol to mitochondria in response to either NU14 or purified FimH. These data suggest that FimH acts as a tethered toxin of UPEC that activates caspase-dependent urothelial apoptosis via direct induction of the extrinsic pathway and that the intrinsic pathway is activated indirectly as a result of coupling by caspase 8-mediated Bid cleavage.
authors.affiliation: Department of Urology, Feinberg School of Medicine, Northwestern University, 303 East Chicago Avenue, Chicago, IL 60611, USA. d-klumpp@northwestern.edu
authors.forename: David J
authors.initials: DJ
authors.lastname: Klumpp
authors.forename: Matthew T
authors.initials: MT
authors.lastname: Rycyk
authors.forename: Michael C
authors.initials: MC
authors.lastname: Chen
authors.forename: Praveen
authors.initials: P
authors.lastname: Thumbikat
authors.forename: Shomit
authors.initials: S
authors.lastname: Sengupta
authors.forename: Anthony J
authors.initials: AJ
authors.lastname: Schaeffer
mesh_terms: D018830:Adhesins, Escherichia coli / Q000378:metabolism*; D017209:Apoptosis; D051023:BH3 Interacting Domain Death Agonist Protein / Q000378:metabolism; D001422:Bacterial Adhesion; D002118:Calcium / Q000378:metabolism; D053143:Caspase 2; D053181:Caspase 8; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004926:Escherichia coli / Q000378:metabolism / Q000472:pathogenicity*; D004927:Escherichia coli Infections / Q000382:microbiology* / Q000473:pathology; D036781:Fimbriae Proteins / Q000378:metabolism*; D005434:Flow Cytometry; D006801:Humans; D051336:Mitochondrial Membranes / Q000378:metabolism; D008954:Models, Biological; D021381:Protein Transport; D014552:Urinary Tract Infections / Q000382:microbiology* / Q000473:pathology; D019459:Urothelium / Q000382:microbiology* / Q000473:pathology; D051020:bcl-X Protein / Q000378:metabolism
chemical_list: D018830:Adhesins, Escherichia coli; C494811:BCL2L1 protein, human; D051023:BH3 Interacting Domain Death Agonist Protein; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D051020:bcl-X Protein; C058769:fimH protein, E coli; D036781:Fimbriae Proteins; C505530:CASP8 protein, human; D053143:Caspase 2; D053181:Caspase 8; D020169:Caspases; D002118:Calcium