Monocyte 15-lipoxygenase expression is regulated by a novel cytosolic signaling complex with protein kinase C δ and tyrosine-phosphorylated Stat3

Our previous studies demonstrated that the IL-13-induced 15-lipoxygenase expression in primary human monocytes is regulated by the activation of both Stat1 and Stat3 and by protein kinase C (PKC)delta. IL-13 stimulated the phosphorylation of Stat3 on both Tyr(705) and Ser(727). In this study we show that IL-13 induces the association of PKCS with Stat3, not with Stall, and is required for Stat3 Ser 727 phosphorylation. We found a novel IL-13-dependent cytosolic signaling complex of PKCS and tyrosine-phosphorylated Stat3. A tyrosine kinase inhibitor blocked PKC delta association with Stat3 as well as Stat3 Ser 727 phosphorylation. We therefore hypothesized that tyrosine phosphorylation was required for Stat3 interaction with PKC delta and subsequent PKC delta-5-dependent phosphorylation of Stat3 Ser(727). We developed an efficient transfection protocol for human monocytes. Expression of Stat3 containing a mutation in Tyr(705) inhibited the association of PKC5 with Stat3 and blocked Stat3 Ser 727 phosphorylation, whereas transfection with wild-type Stat3 did not. Furthermore, by transfecting monocytes with Stat3 containing mutations in Tyr(705) or Ser(727) or with wild-type Stat3, we demonstrated that both Stat3 tyrosine and serine phosphorylations are required for optimal binding of Stat3 with DNA and maximal expression of 15-lipoxygenase, an important regulator of inflammation and apoptosis.