title: Regulation of Rac1 and Reactive Oxygen Species Production in Response to Infection of Gastrointestinal Epithelia.
abstract: Generation of reactive oxygen species (ROS) during infection is an immediate host defense leading to microbial killing. APE1 is a multifunctional protein induced by ROS and after induction, protects against ROS-mediated DNA damage. Rac1 and NAPDH oxidase (Nox1) are important contributors of ROS generation following infection and associated with gastrointestinal epithelial injury. The purpose of this study was to determine if APE1 regulates the function of Rac1 and Nox1 during oxidative stress. Gastric or colonic epithelial cells (wild-type or with suppressed APE1) were infected with Helicobacter pylori or Salmonella enterica and assessed for Rac1 and NADPH oxidase-dependent superoxide production. Rac1 and APE1 interactions were measured by co-immunoprecipitation, confocal microscopy and proximity ligation assay (PLA) in cell lines or in biopsy specimens. Significantly greater levels of ROS were produced by APE1-deficient human gastric and colonic cell lines and primary gastric epithelial cells compared to control cells after infection with either gastric or enteric pathogens. H. pylori activated Rac1 and Nox1 in all cell types, but activation was higher in APE1 suppressed cells. APE1 overexpression decreased H. pylori-induced ROS generation, Rac1 activation, and Nox1 expression. We determined that the effects of APE1 were mediated through its N-terminal lysine residues interacting with Rac1, leading to inhibition of Nox1 expression and ROS generation. APE1 is a negative regulator of oxidative stress in the gastrointestinal epithelium during bacterial infection by modulating Rac1 and Nox1. Our results implicate APE1 in novel molecular interactions that regulate early stress responses elicited by microbial infections.
authors.affiliation: Department of Medicine, University of California, San Diego, La Jolla, California, United States of America.
authors.forename: Gerco
authors.initials: G
authors.lastname: den Hartog
authors.affiliation: Department of Medicine, University of California, San Diego, La Jolla, California, United States of America.
authors.forename: Ranajoy
authors.initials: R
authors.lastname: Chattopadhyay
authors.affiliation: Department of Pathology, University of California, San Diego, La Jolla, California, United States of America.
authors.forename: Amber
authors.initials: A
authors.lastname: Ablack
authors.affiliation: Department of Surgery, University of Virginia, Charlottesville, Virginia, United States of America.
authors.forename: Emily H
authors.initials: EH
authors.lastname: Hall
authors.affiliation: Department of Medicine, University of California, San Diego, La Jolla, California, United States of America.
authors.forename: Lindsay D
authors.initials: LD
authors.lastname: Butcher
authors.affiliation: National Institute of Science Education and Research (NISER), Bhubaneswar, India.
authors.forename: Asima
authors.initials: A
authors.lastname: Bhattacharyya
authors.affiliation: Department of Medicine, University of California, San Diego, La Jolla, California, United States of America.
authors.forename: Lars
authors.initials: L
authors.lastname: Eckmann
authors.affiliation: Division of Pediatrics, Unit of Gastroenterology and Nutrition, School of Medicine, Pontifical Catholic University, Santiago, Chile.
authors.forename: Paul R
authors.initials: PR
authors.lastname: Harris
authors.affiliation: Department of Pathology, University of California, San Diego, La Jolla, California, United States of America.
authors.forename: Soumita
authors.initials: S
authors.lastname: Das
authors.affiliation: Department of Pathology, University of California, San Diego, La Jolla, California, United States of America.
authors.forename: Peter B
authors.initials: PB
authors.lastname: Ernst
authors.affiliation: Department of Medicine, University of California, San Diego, La Jolla, California, United States of America.
authors.forename: Sheila E
authors.initials: SE
authors.lastname: Crowe
mesh_terms: D015153:Blotting, Western; D002460:Cell Line; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D005753:Gastric Mucosa / Q000276:immunology* / Q000378:metabolism / Q000382:microbiology; D016481:Helicobacter Infections / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D007413:Intestinal Mucosa / Q000276:immunology* / Q000378:metabolism / Q000382:microbiology; D018613:Microscopy, Confocal; D017382:Reactive Oxygen Species / Q000276:immunology / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D012480:Salmonella Infections / Q000276:immunology* / Q000378:metabolism; D020830:rac1 GTP-Binding Protein / Q000276:immunology / Q000378:metabolism*
chemical_list: C107953:RAC1 protein, human; D017382:Reactive Oxygen Species; D020830:rac1 GTP-Binding Protein; C085108:APEX1 protein, human; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase