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.

Human and Helicobacter pylori Interactions Determine the Outcome of Gastric Diseases

“…This provides a rationale for the use of NOX1 as a marker of neoplastic transformation and for the role of NOX-1-derived ROS in H. pylori -mediated carcinogenesis in the human stomach. In the same way, non-transformed antral-derived primary epithelial cells and the gastric epithelial cell line AGS infected with H. pylori 26695 express NOX1 and produce ROS rapidly (within 30 min) and transiently (den Hartog et al 2016). The transcription factor(s) involved in H. pylori -mediated NOX1 transcription in gastric epithelial cells have not been characterized yet, but one study implicated the pleiotropic signaling molecule Ras-related C3 botulinum toxin substrate 1 (RAC1) (den Hartog et al 2016).…”

“…In the same way, non-transformed antral-derived primary epithelial cells and the gastric epithelial cell line AGS infected with H. pylori 26695 express NOX1 and produce ROS rapidly (within 30 min) and transiently (den Hartog et al 2016). The transcription factor(s) involved in H. pylori -mediated NOX1 transcription in gastric epithelial cells have not been characterized yet, but one study implicated the pleiotropic signaling molecule Ras-related C3 botulinum toxin substrate 1 (RAC1) (den Hartog et al 2016). Further work is warranted to delineate the molecular mechanisms leading to NOX1 expression, since these findings could provide critical insight into H. pylori -associated inflammation.…”

“…Lastly, α-lipoic acid, a naturally occurring thiol compound that exhibits antioxidant properties, inhibits NADPH oxidase-derived ROS production in AGS cells and concomitantly dampens NF-κB and AP-1 activation and expression of the oncogenes β-catenin (CTNNB1) and c-Myc (MYC) (Byun et al 2014). Interestingly, the endonuclease apurinic/apyrimidinic endodeoxyribonuclease 1 (APEX1, also known as APE1), a multifunctional enzyme that plays a central role in the cellular response to oxidative stress, including DNA repair and redox regulation of transcriptional factors, is induced by endogenous ROS in AGS cells (den Hartog et al 2016). APEX1 inhibits RAC1 activation and consequently NOX1-dependent ROS production, and blocking APEX1 leads to sustained NOX1 activation and increased O 2 − generation (den Hartog et al 2016).…”

“…Interestingly, the endonuclease apurinic/apyrimidinic endodeoxyribonuclease 1 (APEX1, also known as APE1), a multifunctional enzyme that plays a central role in the cellular response to oxidative stress, including DNA repair and redox regulation of transcriptional factors, is induced by endogenous ROS in AGS cells (den Hartog et al 2016). APEX1 inhibits RAC1 activation and consequently NOX1-dependent ROS production, and blocking APEX1 leads to sustained NOX1 activation and increased O 2 − generation (den Hartog et al 2016). This could represent a mechanism by which cells limit NOX1-dependent DNA damage and/or a strategy developed by the bacterium, to promote its own survival by inhibiting ROS production.…”

Polyamine- and NADPH-dependent generation of ROS during Helicobacter pylori infection: A blessing in disguise

“…This suggests that NOX1 could represent a marker of neoplastic transformation. In addition, non-transformed antral-derived primary epithelial cells and the gastric epithelial cell line AGS infected with H. pylori 26695 express NOX1 through a signal transduction process involving Ras-related C3 botulinum toxin substrate 1 (RAC1), which is also a protein partner for NOX1, and produce ROS rapidly and transiently [56]. Similarly, NOX1 induction and ROS production has been observed in primary guinea pig gastric mucosal cells infected with H. pylori lipopolysaccharide (LPS) [57] through a PI3K/RAC1-dependent pathway [58].…”