The Ausubel lab uses genetic and whole genome approaches to investigate the molecular basis of microbial pathogenesis and host innate immunity.
The vertebrate innate immune system serves as the first line of defense against microbial invaders, and activates the other branch of immunity, the adaptive immune system. Although the adaptive immune system, involving the clonal selection of lymphocytes, is unique to vertebrates, the innate immune response has ancient origins. Innate immunity is characterized by receptors that are activated by signals indicative of pathogens, often referred to as pathogen-associated molecular patterns (PAMPs). PAMPs include conserved microbial cell wall molecules such as lipopolysaccharide and peptidoglycan. PAMP receptors activate downstream signaling cascades; ultimately, a set of immune effectors is activated.
Comprehensive genetic analyses of the molecules that comprise the innate immune response have only recently been initiated. Model systems, including Drosophila melanogaster, have proven invaluable in the identification of new components of innate immunity, many of which have conserved roles in vertebrates. Numerous studies now suggest that the fundamental mechanisms of host defense against pathogen invasion are conserved across metazoans, highlighting the importance of continued genetic analysis of the immune system in model species.
The Ausubel lab has pioneered the use of the nematode Caenorhabditis elegans as a new model host to study the innate immune response to diverse pathogens. This nematode-pathogen model system is advantageous because worms are simply fed various bacteria or yeast in place of their usual food source of Escherichia coli (strain OP50) and observed for their survival over time. A remarkably large number of human pathogens have been found to be lethal to C. elegans in this assay, including the Gram-positive bacteria Staphyloccus aureus and Enterococcus faecalis, the Gram-negative bacteria Pseudomonas aeruginosa, and Salmonella enterica, and the yeasts Cryptococcus neoformans and Candida albicans.
Powerful biochemical, molecular, and forward and reverse genetic tools are readily available in C. elegans, permitting comprehensive analysis of both bacterial virulence factors and host defense factors. Both pathogen and host immune molecules have been identified by the Ausubel lab and others that overlap significantly with known factors in mammalian models, indicating the evolutionary conservation of pathogenesis and host immune response and the relevance of the nematode model system to vertebrates. Additionally, novel bacterial genes with previously unknown roles in pathogenesis have been discovered using the C. elegans model system and subsequently shown to be required for pathogenesis in mice. Thus, the C. elegans-pathogen system has the potential for the rapid identification and characterization of the signaling molecules and complex pathways that comprise the innate immune response and the bacterial factors that function in the pathogenesis process.
Using both forward and reverse genetic approaches, we have identified at least six C. elegans genes (tir-1, nsy-1, sek-1, pmk-1, mek-1, and vhp-1) that function in a conserved p38 MAPK immune signaling pathway. tir-1 encodes a TIR-domain-containing protein that functions upstream of a MAPK cascade encoded by nsy-1 (MAPKKK), sek-1 (MAPKK) and pmk-1 (p38 MAPK). NSY-1 and SEK-1 are required for PMK-1 activation and the MAPKK encoded by mek-1 enhances PMK-1 (p38) phosphorylation. vhp-1 encodes a MAPK phosphatase and appears to work in conjunction with SEK-1 and MEK-1 to fine tune levels of PMK-1 activation. We have also shown that long-lived mutants in the DAF-2 insulin-like signaling pathway are highly resistant to both Gram-positive and Gram-negative pathogens and that this resistance is suppressed by mutations in the p38 MAPK pathway. Our microarray studies of the genes regulated by the p38 MAPK pathway and the DAF-2-DAF-16 pathway suggest that the p38 MAPK pathway acts in parallel to DAF-2-DAF-16 pathway to regulate immune gene expression.