Ansfected with L. monocytogenes RNA, L. monocytogenes DNA or 3PdsRNA. Kind I IFN (THP-1, A549, Colo205) or CXCL10 (HepG2) production was analyzed 24 hours following stimulation. The relative induction from the indicated cytokine is depicted as percentage of induction by transfected L. monocytogenes (L.M.) RNA. C, D, E and F: THP-1, A549, HepG2 and Colo205 cells have been infected with wt and hly- L. monocytogenes at the indicated MOI. Kind I IFN (THP-1, A549, Colo205) or CXCL10 production (HepG2) was analyzed 24 hours after stimulation. Error bars represent s.d. doi:ten.1371/journal.pone.0062872.gnot raise a form I IFN response upon transfection of bacDNA, in spite of a robust response to poly(dA-dT), indicating the absence of functional STING-dependent recognition pathways. The latter acquiring also excludes an involvement of your pol III/RIG-I dependent pathway of bacDNA detection as suggested by Chu et al. [24] but corroborates information of Monroe et al. [59] for L. pneumophila bacterial DNA, which was shown not to trigger the pol III/RIG-I dependent pathway. Regardless of translocation of bacRNA into the cytosol, RNAi of RIG-I or MAVS in monocytic cells (THP-1) didn’t impair kind I IFN induction through infection with L. monocytogenes. This suggests a redundant part of RIG-I regarding form I IFN induction inmonocytes throughout L. monocytogenes infection, as shown for L. pneumophila [59]. The minor contribution on the RIG-I/MAVS pathway in monocytic cells may be resulting from a dominance in concentration or impact of other stimuli released by L. monocytogenes, like bacDNA or the lately discovered mediator cyclic diadenosine monophosphate (c-di-AMP) [34]. By contrast, induction of kind I IFN in the course of L. monocytogenes infection of epithelial cells was practically completely abolished by RNAi-mediated knockdown of RIG-I or MAVS, implicating that neither DNA nor c-di-AMP plays a function for L. monocytogenes mediated sort I IFN induction in these cells. With each other, we conclude that RIG-I plays a major part inside the L. monocytogenes-PLOS A single | www.plosone.orgRIG-I Detects RNA of Listeria in Non-Immune CellsFigure four. Knockdown of RIG-I abrogates L. monocytogenes-induced kind I IFN induction in epithelial but not in monocytic cells. A: Murine BM-DCs were transfected with indicated stimuli. 1 out of two experiments is shown. Murine IFN-a secretion was analyzed 24 hours after stimulation.Anti-Mouse IL-10 Antibody Error bars represent SEM.Penciclovir B: A549 cells had been transfected with siRNA against RIG-I, MAVS or Luciferase (control).PMID:22943596 Cells had been then infected with L. monocytogenes or transfected with L. monocytogenes RNA (L.m. RNA), L. monocytogenes DNA (L.m. RNA) or 3P-dsRNA 48 hours immediately after knockdown. Form I IFN production was analyzed 24 hours right after stimulation. B) THP-1 cells have been electroporated with control siRNA or siRNAs against MAVS or STING. 72 hours right after electroporation, THP-1 cells have been infected with hly- or wt L. monocytogenes at indicated MOI or transfected with plasmid DNA (pDNA) or RIG-I ligand (3P-dsRNA). Form I IFN production was analyzed 24 hours soon after stimulation. Relative form I IFN induction was normalized to cells transfected with siRNA against Luciferase and stimulated with pDNA. Error bars represent s.d. doi:ten.1371/journal.pone.0062872.gPLOS One | www.plosone.orgRIG-I Detects RNA of Listeria in Non-Immune Cellsinduced secretion of type-I-IFN in cell varieties lacking functional STING-dependent pathways. Cytosolic recognition of bacterial RNA is controversial. Mancuso and colleagues [60] showed that phag.
