Pathogens subvert the host immune response by inhibiting microbiota-mediated activation of the aryl hydrocarbon receptor

Author(s):
Rebecca Meltzer; Robert Keskey; Anne Dijkstra; Sanjiv Hyoju; Ashley Sidebottom; Jessica Little; Abhimanyu Thakur; Kui Zhang; Huanhuan Joyce Chen; Olga Zaborina; John Alverdy

Background:

The success of a pathogen is a function of its ability to subvert the host immune system. We have demonstrated that the gut microbiota plays a key role in host clearance of invading pathogens by metabolizing tryptophan into indoles, which activate host macrophages via the aryl hydrocarbon receptor (AhR). We further found that pathogens secrete products that can interfere with the AhR-directed immune response. However, the mechanism of pathogen immune subversion and molecular details of this interaction remain unclear.

Hypothesis:

We hypothesized that exoproducts secreted by pathogens can compete with gut microbiota-derived indoles to bind to the aryl hydrocarbon receptor (AhR) on host macrophages and increase mortality from lethal bacterial infection.

Methods:

Common virulence factors and their interactions with AhR were modeled in silico utilizing molecular docking (AutoDock) and matched with whole genome sequencing and metabolomics of Serratia marcescens (Sm) and Klebsiella oxytoca (Ko) exoproducts. The ability of identified molecules to suppress indole activation of AhR was studied in vitro utilizing a murine AhR reporter cell line. Finally, in vivo studies were performed using a murine model of lethal bacterial infection with intraperitoneal (IP) Sm to determine if co-administration of candidate AhR inhibiting molecules would increase mortality in mice.

Results:

Supernatants from clinically relevant pathogens (i.e., Sm, Ko, or a polymicrobial community) demonstrated a conserved ability to inhibit indole activation of AhR in vitro. In silico modeling of AhR interaction with indoles and low molecular weight pathogen-secreted virulence factors was compared with whole genome sequencing and metabolomics of Sm and Ko, which identified enterobactin among microbial exoproducts capable of binding to AhR. In silico, enterobactin was found to have a more stable bond to AhR than the main AhR-binding indole molecule, indole-3-acetic acid, implicating enterobactin’s ability to compete with indole binding to AhR. In vitro, enterobactin significantly suppressed indole-mediated activation of AhR by 89%. In vivo, co-administration of enterobactin with IP Sm significantly increased mortality (73.3% vs 26.7%, n=15/group, p=0.016).

Conclusions:

Pathogen-derived virulence factors can compete with gut microbiota-derived indole metabolites for host aryl hydrocarbon receptor signaling, identifying a novel mechanism of pathogen subversion of the host immune response to increase mortality from lethal peritoneal infection.