Dafachronic acid and temperature regulate canonical dauer pathways during Nippostrongylus brasiliensis infectious larvae activation

Background: While immune responses to the murine hookworm Nippostrongylus brasiliensis have been investigated, signaling pathways regulating development of infectious larvae (iL3) are not well understood. We hypothesized that N. brasiliensis would use pathways similar to those controlling dauer development in the free-living nematode Caenorhabditis elegans, which is formally known as the "dauer hypothesis." Methods: To investigate whether dafachronic acid activates the N. brasiliensis DAF-12 homolog, we utilized an in vitro reporter assay. We then utilized RNA-Seq and subsequent bioinformatic analyses to identify N. brasiliensis dauer pathway homologs and examine regulation of these genes during iL3 activation. Results: In this study, we demonstrated that dafachronic acid activates the N. brasiliensis DAF-12 homolog. We then identified N. brasiliensis homologs for members in each of the four canonical dauer pathways and examined their regulation during iL3 activation by either temperature or dafachronic acid. Similar to C. elegans, we found that transcripts encoding antagonistic insulin-like peptides were significantly downregulated during iL3 activation, and that a transcript encoding a phylogenetic homolog of DAF-9 increased during iL3 activation, suggesting that both increased insulin-like and DAF-12 nuclear hormone receptor signaling accompanies iL3 activation. In contrast to C. elegans, we observed a significant decrease in transcripts encoding the dauer transforming growth factor beta ligand DAF-7 during iL3 activation, suggesting a different role for this pathway in parasitic nematode development. Conclusions: Our data suggest that canonical dauer pathways indeed regulate iL3 activation in the hookworm N. brasiliensis and that DAF-12 may be a therapeutic target in hookworm infections.[Figure not available: see fulltext.]