title: Escherichia coli virulence protein NleH1 interaction with the v-Crk sarcoma virus CT10 oncogene-like protein (CRKL) governs NleH1 inhibition of the ribosomal protein S3 (RPS3)/nuclear factor κB (NF-κB) pathway.
abstract: Enterohemorrhagic Escherichia coli and other attaching/effacing bacterial pathogens cause diarrhea in humans. These pathogens use a type III secretion system to inject virulence proteins (effectors) into host cells, some of which inhibit the innate immune system. The enterohemorrhagic E. coli NleH1 effector prevents the nuclear translocation of RPS3 (ribosomal protein S3) to inhibit its participation as a nuclear "specifier" of NF-κB binding to target gene promoters. NleH1 binds to RPS3 and inhibits its phosphorylation on Ser-209 by IκB kinase-β (IKKβ). However, the precise mechanism of this inhibition is unclear. NleH1 possesses a Ser/Thr protein kinase activity that is essential both for its ability to inhibit the RPS3/NF-κB pathway and for full virulence of the attaching/effacing mouse pathogen Citrobacter rodentium. However, neither RPS3 nor IKKβ is a substrate of NleH1 kinase activity. We therefore screened ∼9,000 human proteins to identify NleH1 kinase substrates and identified CRKL (v-Crk sarcoma virus CT10 oncogene-like protein), a substrate of the BCR/ABL kinase. Knockdown of CRKL abundance prevented NleH1 from inhibiting RPS3 nuclear translocation and NF-κB activity. CRKL residues Tyr-198 and Tyr-207 were required for interaction with NleH1. Lys-159, the kinase-active site of NleH1, was necessary for its interaction with CRKL. We also identified CRKL as an IKKβ interaction partner, mediated by CRKL Tyr-198. We propose that the CRKL interaction with IKKβ recruits NleH1 to the IKKβ complex, where NleH1 then inhibits the RPS3/NF-κB pathway.
authors.affiliation: From the College of Veterinary Medicine, Kansas State University, Manhattan, Kansas 66506 and.
authors.forename: Thanh H
authors.initials: TH
authors.lastname: Pham
authors.forename: Xiaofei
authors.initials: X
authors.lastname: Gao
authors.forename: Gyanendra
authors.initials: G
authors.lastname: Singh
authors.forename: Philip R
authors.initials: PR
authors.lastname: Hardwidge
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism / Q000472:pathogenicity*; D004927:Escherichia coli Infections / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D057809:HEK293 Cells; D054884:Host-Pathogen Interactions / Q000235:genetics*; D006801:Humans; D051550:I-kappa B Kinase / Q000235:genetics / Q000378:metabolism; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D012269:Ribosomal Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C082869:CRKL protein; D029968:Escherichia coli Proteins; D016328:NF-kappa B; C556114:NleH protein, E coli; D009687:Nuclear Proteins; C578290:RPS3 protein, human; D012269:Ribosomal Proteins; D051550:I-kappa B Kinase
keywords: CRKL; Escherichia coli; IKKβ; Innate Immunity; NF-κB; NleH1; Type III Secretion System; Virulence factors