The Evolutionarily Conserved Mediator Subunit MDT-15/MED15 Links Protective Innate Immune Responses and Xenobiotic Detoxification

Ausubel FM (2014) The Evolutionarily Conserved Mediator Subunit MDT-15/MED15 Links Protective Innate Immune Responses and Xenobiotic Detoxification. PLoS Pathog 10(5): e1004143. doi:10.1371/journal.ppat.1004143 The Evolutionarily Conserved Mediator Subunit MDT-15/ MED15 Links Protective Innate Immune Responses and Xenobiotic Detoxification Read Pukkila-Worley 0 Rhonda L. Feinbaum 0 Deborah L. McEwan 0 Annie L. Conery 0 Frederick M. Ausubel 0 Robin Charles May, University of Birmingham, United Kingdom 0 1 Division of Infectious Diseases; Massachusetts General Hospital; Harvard Medical School; Boston, Massachusetts, United States of America, 2 Department of Molecular Biology; Massachusetts General Hospital; Harvard Medical School; Boston, Massachusetts, United States of America, 3 Department of Genetics; Harvard Medical School; Boston , Massachusetts , United States of America Metazoans protect themselves from environmental toxins and virulent pathogens through detoxification and immune responses. We previously identified a small molecule xenobiotic toxin that extends survival of Caenorhabditis elegans infected with human bacterial pathogens by activating the conserved p38 MAP kinase PMK-1 host defense pathway. Here we investigate the cellular mechanisms that couple activation of a detoxification response to innate immunity. From an RNAi screen of 1,420 genes expressed in the C. elegans intestine, we identified the conserved Mediator subunit MDT-15/ MED15 and 28 other gene inactivations that abrogate the induction of PMK-1-dependent immune effectors by this small molecule. We demonstrate that MDT-15/MED15 is required for the xenobiotic-induced expression of p38 MAP kinase PMK1-dependent immune genes and protection from Pseudomonas aeruginosa infection. We also show that MDT-15 controls the induction of detoxification genes and functions to protect the host from bacteria-derived phenazine toxins. These data define a central role for MDT-15/MED15 in the coordination of xenobiotic detoxification and innate immune responses. - Funding: This study was supported by the Cancer Research Institute-Irvington Institute Fellowship Program (to RPW) and by National Institutes of Health awards K08 AI081747 (to RPW), F32 AI098307 (to DLM), R01 AI085581 (to FMA), and P01 AI083214 (to FMA). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. . These authors contributed equally to this work. In nature, organisms encounter environmental insults, such as chemical toxins, secreted microbial virulence factors and invasive pathogens, that threaten their ability to survive and reproduce. As a result, metazoans have evolved protective pathways to counter these challenges. For example, gene families such as cytochrome P450s (CYPs), glutathione-stransferases (GSTs), and UDP-glucuronosyltransferases (UDPs) detoxify xenobiotic small molecule toxins and are conserved from nematodes to humans [1]. Likewise, innate immune defenses provide protection from invasive pathogens [2]. Recent publications have suggested that recognition of xenobiotic toxins is involved in the activation of immune response pathways [3,4]. From an evolutionary perspective, it is logical that hosts respond to threats encountered in the wild at least in part through surveillance pathways that monitor the integrity of core cellular machinery, which are often the targets of xenobiotic small molecules or microbe-generated toxins. These studies predict that organisms may integrate detoxification and immune responses as a means to respond rapidly to such challenges, but the mechanisms underlying this coordinated host response have not been reported. Our research group and others use bacterial and fungal pathogenesis assays in the nematode Caenorhabditis elegans to investigate mechanisms of immune pathway activation in intestinal epithelial cells [2]. Genetic analyses of C. elegans that are hypersusceptible to bacterial infection have revealed that the nematode mounts defense responses through evolutionarily conserved innate immune pathways. For example, the C. elegans NSY-1/SEK-1/PMK-1 Mitogen Activated Protein (MAP) kinase pathway, orthologous to the ASK1 (MAP kinase kinase kinase)/ MKK3/6 (MAP kinase kinase)/p38 (MAP kinase) pathway in mammals, is required for protection against pathogens [5]. C. elegans animals carrying loss-of-function mutations in this pathway have defects in the basal and pathogen-induced expression of immune effectors and are hypersusceptible to killing by bacterial and fungal pathogens [57]. We previously used a C. elegans pathogenesis assay as a means to identify small molecules that protect the host during bacterial infection [8]. One of the compounds identified in this screen, a small molecule called RPW-24, extended the survival of nematodes infected with the human bacterial pathogen Pseudomonas aeruginosa by stimulating the host immune response via the p38 MAP kinase PMK-1 pathway [9]. A genome-wide microarray analysis of animals exposed to RPW-24 revealed that, in addition Metazoans respond to environmental threats in part through conserved pathways that coordinate protective transcriptional responses. During infection with an invasive pathogen, for example, innate immune pathways regulate the secretion of antimicrobial immune effectors. Likewise, exposure to toxic molecules leads to the induction of detoxification mechanisms that protect the host from the deleterious effects of these compounds. Here we find that a conserved transcriptional regulator MDT-15/MED15 links xenobiotic detoxification and immune responses in a manner that is important for protection during bacterial infection. We also show that MDT-15/MED15 is necessary for the host to resist the lethal effects of secreted toxins produced by pathogenic bacteria. Rapid coordination of these protective host responses through MDT-15/MED15 may therefore be part of a conserved survival strategy in the wild. to inducing the transcription of putative immune effectors, this molecule also strongly upregulated Phase I and Phase II detoxification enzymes (CYPs, GSTs and UDPs), suggesting that RPW-24 is a xenobiotic toxin to C. elegans. Consistent with this hypothesis, RPW-24 caused a dose dependent reduction of nematode lifespan on nonpathogenic food and delayed development of animals that were exposed starting at the first larval stage. Here we sought to use RPW-24 as a tool to characterize mechanisms of p38 MAP kinase PMK-1 pathway activation in C. elegans. We found that activation of PMK-1-regulated pathogen response genes is genetically linked to the induction of genes involved in the detoxification of small molecule toxins. We show that the evolutionarily conserved Mediator subunit MDT-15/MED15 is required for the induction of the p38 MAP kinase PMK-1-mediated immune effectors as well as (...truncated)