Peroxidation of polyunsaturated fatty acids leads to the formation of a large array of lipid-derived electrophiles (LDEs), many of which are important signaling molecules involved in the pathogenesis of human diseases. Previous research has shown that one of such LDEs, trans, trans-2,4-decadienal (tt-DDE), increases inflammation, however, the underlying mechanisms are not well understood. Here we used click chemistry-based proteomics to identify the cellular targets which are required for the pro-inflammatory effects of tt-DDE. We found that treatment with tt-DDE increased cytokine production, JNK phosphorylation, and activation of NF-?B signaling in macrophage cells, and increased severity of dextran sulfate sodium (DSS)-induced colonic inflammation in mice, demonstrating its pro-inflammatory effects in vitro and in vivo. Using click chemistry-based proteomics, we found that tt-DDE directly interacts with Hsp90 and 14-3-3?, which are two important proteins involved in inflammation and tumorigenesis. Furthermore, siRNA knockdown of Hsp90 or 14-3-3? abolished the pro-inflammatory effects of tt-DDE in macrophage cells. Together, our results support that tt-DDE increases inflammatory responses via Hsp90- and 14-3-3?-dependent mechanisms.
Systemic RNA interference (RNAi) in Caenorhbaditis elegans requires sid-1, sid-3, and sid-5 Injected, expressed, or ingested double-stranded RNA (dsRNA) is transported between cells, enabling RNAi in most tissues, including the germline and progeny (parental RNAi). A recent report claims that parental RNAi also requires the yolk receptor rme-2 Here, we characterize the role of the sid genes and rme-2 in parental RNAi. We identify multiple independent paths for maternal dsRNA to reach embryos and initiate RNAi. We showed previously that maternal and embryonic sid-1 contribute independently to parental RNAi. Here we demonstrate a role for embryonic sid-5, but not sid-2 or sid-3 in parental RNAi. We also find that maternal rme-2 contributes to but is not required for parental RNAi. We determine that parental RNAi by feeding occurs nearly exclusively in adults. We also introduce 5-ethynyluridine to densely internally label dsRNA, avoiding complications associated with other labeling strategies such as inhibition of normal dsRNA trafficking and separation of label and RNA. Labeling shows that yolk and dsRNA do not colocalize following endocytosis, suggesting independent uptake, and, furthermore, dsRNA appears to rapidly progress through the RAB-7 endocytosis pathway independently of sid-1 activity. Our results support the premise that although sid-1 functions in multiple roles, it alone is central and absolutely required for inheritance of silencing RNAs.
