Engagement of Toll-like receptor (TLR) proteins activates multiple signal transduction pathways. Previous studies demonstrated that TLR2 and TLR4 engagement leads to rapid phosphorylation of the transcription factor STAT1 at serine 727 (Ser-727 STAT1) in murine macrophages. Only TLR4 engagement induced STAT1 phosphorylation at tyrosine 701, although this response was delayed compared with Ser-727 STAT1 phosphorylation. Unlike other cell types, the p38 mitogen-activated protein kinase was necessary, but not sufficient, for TLR-induced phosphorylation of Ser-727 STAT1 in macrophages. We and others had previously shown that Ser-727 STAT1 phosphorylation could be blocked by rottlerin, an inhibitor of protein kinase C-δ (PKC—δ). Here we report that peritoneal exudate macrophages from PKC-δ-deficient mice can be activated through TLR2 and TLR4 to elicit rapid phosphorylation of Ser-727 STAT1, which was blocked by both rottlerin and the p38 inhibitor SB203580, but not by the pan-PKC inhibitor bisindoylmaleamide. Furthermore, both normal and PKC-δ-deficient macrophages secreted comparable amounts of IL-6, IP-10, and RANTES following TLR engagement. In contrast, IFN-γ-induced STAT1 serine phosphorylation was independent of both PKC-δ and p38. Overall, these studies demonstrate that a PKC-δindependent signaling pathway downstream of both TLR2 and TLR4 is necessary for Ser-727 STAT1 phosphorylation in primary murine macrophages.
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