Tunicamycin inhibits Toll-like receptor-activated inflammation in RAW264.7 cells by suppression of NF-κB and c-Jun activity via a mechanism that is independent of ER-stress and N-glycosylation.
In this study, we investigated the effect of tunicamycin on the production of pro-inflammatory molecules in RAW264.7 macrophage cells in response to lipopolysaccharide (LPS) and Toll-like receptor (TLR) agonists. Tunicamycin caused a reduction in LPS-induced nitric oxide (NO) production and expression of inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), interleukin-1 beta (IL-1β) and tumor necrosis factor alpha (TNF-α). In contrast, other ER stress-inducing chemicals, such as A23187 and thapsigargin (TG), increased LPS-induced COX-2 expression and had no effect on LPS-induced iNOS, TNF-α or IL-1β expression. Furthermore, the inhibitory effect of tunicamycin on LPS-induced inflammation was not influenced by salubrinal, an ER stress inhibitor, suggesting that the anti-inflammatory effect of tunicamycin is independent of ER stress. Tunicamycin also inhibited the expression of inflammatory molecule mRNAs induced by stimulation of TLR2 (with lipoteichoic acid) or TLR3 (with polyinosinic:polycytidylic acid), which do not require myeloid differentiation protein-2 (MD2) for their activation. Moreover, inhibition of LPS-induced iNOS expression was not inhibited by castanospermine, another N-glycosylation inhibitor, suggesting that the inhibitory effect of tunicamycin on LPS-induced iNOS induction is likely independent of MD2 N-glycosylation. Tunicamycin inhibited nuclear factor-kappaB (NF-κB) activity by suppressing LPS-induced nuclear translocation of p50 and subsequent DNA binding of p50 and p65 to the NF-κB site of the iNOS promoter. Tunicamycin also inhibited the transcriptional activity of a cAMP-response element (CRE) reporter, possibly by inhibiting c-Jun activation. Therefore, we conclude that tunicamycin represses TLR-induced inflammation through suppression of NF-κB and CRE activity via a mechanism that is independent of ER-stress and N-glycosylation.
Department of Physiology and Biophysics, College of Medicine, Inha University, 253, Shinheung-Dong, Jung-Ku, Incheon, Republic of Korea.
Song-Yi
SY
Kim
Ji-Sun
JS
Hwang
Inn-Oc
IO
Han
D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D000818:Animals; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D051546:Cyclooxygenase 2 / Q000235:genetics; D004247:DNA / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D007249:Inflammation / Q000378:metabolism; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D052002:NF-kappa B p50 Subunit / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000235:genetics; D051193:Toll-Like Receptors / Q000378:metabolism*; D051996:Transcription Factor RelA / Q000378:metabolism*; D014415:Tunicamycin / Q000494:pharmacology*; D015854:Up-Regulation / Q000187:drug effects
D017362:Cyclic AMP Response Element-Binding Protein; D008070:Lipopolysaccharides; D052002:NF-kappa B p50 Subunit; D051193:Toll-Like Receptors; D051996:Transcription Factor RelA; D014415:Tunicamycin; D009569:Nitric Oxide; D004247:DNA; D052247:Nitric Oxide Synthase Type II; D051546:Cyclooxygenase 2; D048031:JNK Mitogen-Activated Protein Kinases
ER stress; Inflammation; Tunicamycin