Lack of Tir ubiquitylation contributes to enteropathogenic E. coli remaining extracellular during nonphagocytic cell infections.
Enteropathogenic Escherichia coli (EPEC) is an extracellular pathogen that alters many host subcellular components during its infectious processes. We have previously shown that EPEC hijacks a large assortment of host cell endocytic components and uses these proteins to form protruding structures called "pedestals" rather than triggering internalization of the bacteria. Other invasive pathogens that also recruit similar endocytic components have been shown to enter their host cells on the ubiquitylation of their host cell receptors. Therefore, we hypothesize that EPEC remains extracellular by maintaining its receptor, translocated intimin receptor (Tir), in an unubiquitylated state. Using immunoprecipitation-Western blots, we demonstrate no association of ubiquitin with Tir. To further elucidate the effect Tir ubiquitylation would have on EPEC during their infections, we engineered Tir-ubiquitin fusion constructs, expressed them in host epithelial cells, and infected them with Δtir EPEC. We found these cells induced a significant increase in EPEC invasion as compared with cells that expressed the Tir construct that lacked ubiquitin conjugation. Our results indicate that the lack of EPEC receptor ubiquitylation is a contributing factor that these microbes use to prevent their internalization into epithelial cells.
Department of Biological Sciences, Simon Fraser University, Shrum Science Centre, Burnaby, British Columbia V5A 1S6, Canada.
Ann E
AE
Lin
Julian A
JA
Guttman
D000818:Animals; D015153:Blotting, Western; D019556:COS Cells; D002462:Cell Membrane / Q000378:metabolism*; D002522:Chlorocebus aethiops; D054308:Enteropathogenic Escherichia coli / Q000254:growth & development / Q000378:metabolism* / Q000472:pathogenicity; D004927:Escherichia coli Infections / Q000378:metabolism* / Q000382:microbiology; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006801:Humans; D047468:Immunoprecipitation; D011485:Protein Binding; D021381:Protein Transport; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination
D029968:Escherichia coli Proteins; D011956:Receptors, Cell Surface; C109287:Tir protein, E coli