Previous studies have suggested that heterotrimeric G proteins and tyrosine kinases may be involved in lipopolysaccharide (LPS) signaling events. Signal transduction pathways activated by LPS were examined in human promonocytic THP-1 cells. We hypothesized that Gi proteins and Src tyrosine kinase differentially affect mitogen-activated protein (MAP) kinases (MAPK) and nuclear factor kappa (NF- B) activation. Post-receptor coupling to G i proteins were examined using pertussis toxin (PTx), which inhibits G i receptor-coupling. The involvement of the Src family of tyrosine kinases was examined using the selective Src tyrosine kinase inhibitor pyrazolopyrimidine-2 (PP2). Pretreatment of THP-1 cells with PTx attenuated LPS-induced activation of c-Jun-N-terminal kinase (JNK) and p38 kinase, and production of tumor necrosis factor-alpha (TNF-) and thromboxane B2 (TxB2). Pretreatment with PP2 inhibited TNF- and TxB2 production, but had no effect on p38 kinase or JNK signaling. Therefore, the G i-coupled signaling pathways and Src tyrosine kinase-coupled signaling pathways are necessary for LPS-induced TNF- and TxB2 production, but differ in their effects on MAPK activation. Neither PTx nor PP2 inhibited LPS-induced activation of interleukin receptor activated kinase (IRAK) or inhibitedtranslocation of NF- B. However, PP2 inhibitedLPS-inducedNF-B transactivation of a luciferase reporter gene construct in a concentration-dependent manner. Thus, LPS induction of Src tyrosine kinases may be essential in downstream NF- B transactivation of genes following DNA binding. PTx had no effect on NF- B activation of the reporter construct. These data suggest upstream divergence in signaling through G i pathways leading to MAPK activation and other signaling events leading to I B degradation and NF- B DNA binding.
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