Abstract

We previously have demonstrated that tumor necrosis factor-α (TNF-α) increases prostaglandin endoperoxide synthase-2 (PGHS-2) mRNA accumulation and tyrosine phosphorylation in the fibrosarcoma cell line, MCA-101. Tyrosine kinase inhibitor, genistein, and tyrosine phosphatase inhibitor, phenylarsine oxide (PAO), blocked TNF-α-mediated induction of PGHS-2 mRNA in these cells. Because the PGHS-2 promoter has a nuclear factor-κB (NF-κB) binding motif, which is important for PGHS-2 gene transcription in some cell types, we have evaluated the effects of tyrosine kinase inhibitors and PAO on TNF-α-induced NF-κB activation. TNF-α (1 nM) rapidly induced translocation of NF-κB, an event accompanied by degradation of inhibitory protein IκB-α. N-tosyl-l-phenylalanine chloromethyl ketone (TPCK), a serine protease inhibitor, inhibited IκB-α degradation and NF-κB activation in response to TNF-α in a dose-dependent manner (25, 50, 100 μM). TPCK also inhibited PGHS-2 mRNA accumulation. These data suggest that NF-κB contributed to PGHS-2 mRNA accumulation in MCA-101 cells stimulated with TNF-α. PAO (2.4 μM) completely abolished activation of NF-κB and degradation of IκB-α induced by TNF-α at a concentration that blocked PGHS-2 mRNA accumulation. However, four tyrosine kinase inhibitors, genistein, tyrphostin 47, herbimycin A and erbstatin, failed to block translocation of NF-κB and degradation of IκB-α. These data demonstrate that tyrosine kinase pathways are not required for TNF-α-induced NF-κB activation in MCA-101 cells and suggest that signaling via these pathways mediates TNF-α-induced PGHS-2 mRNA accumulation via an NF-κB-independent mechanism. Moreover, an upstream tyrosine phosphatase pathway may mediate PGHS-2 mRNA accumulation by TNF-α via an NF-κB-dependent mechanism.