Protein Kinase C ζ Plays a Central Role in Activation of the p42/44 Mitogen-Activated Protein Kinase by Endotoxin in Alveolar Macrophages

Abstract

Human alveolar macrophages respond to endotoxin (LPS) by activation of a number of mitogen-activated protein kinase pathways, including the p42/44 (extracellular signal-related kinase (ERK)) kinase pathway. In this study, we evaluated the role of the atypical protein kinase C (PKC) isoform, PKC ζ, in LPS-induced activation of the ERK kinase pathway. Kinase activity assays showed that LPS activates PKC ζ, mitogen-activated protein/ERK kinase (MEK, the upstream activator of ERK), and ERK. LPS did not activate Raf-1, the classic activator of MEK. Pseudosubstrate-specific peptides with attached myristic acid are cell permeable and can be used to block the activity of specific PKC isoforms in vivo. We found that a peptide specific for PKC ζ partially blocked activation of both MEK and ERK by LPS. We also found that this peptide blocked in vivo phosphorylation of MEK after LPS treatment. In addition, we found that LPS caused PKC ζ to bind to MEK in vivo. These observations suggest that MEK is an LPS-directed target of PKC ζ. PKC ζ has been shown in other systems to be phosphorylated by phosphatidylinositol (PI) 3-kinase-dependent kinase. We found that LPS activates PI 3-kinase and causes the formation of a PKC ζ/PI 3-kinase-dependent kinase complex. These data implicate the PI 3-kinase pathway as an integral part of the LPS-induced PKC ζ activation. Taken as a whole, these studies suggest that LPS activates ERK kinase, in part, through activation of an atypical PKC isoform, PKC ζ.

Phosphatidylinositol 3-Kinase Activity Negatively Regulates Stability of Cyclooxygenase 2 mRNA

“…DISCUSSION We have shown previously that LPS induces production of PGE2 in human alveolar macrophages by increasing amounts of COX2 protein and mRNA (1,5,6). We have also shown that LPS increases the activity of PI3K and its downstream kinase, Akt (13,14). In this study, we show that PI3K activity regulates expression of COX2 and release of PGE2 negatively.…”

Reciprocal Regulation of Protein Kinase C Isoforms Results in Differential Cellular Responsiveness

“…PKCb inhibitor is an anilino-monoindolylmaleimide compound that acts as a potent, cell-permeable inhibitor of PKCb isozymes (25). PKCz/l inhibitor is a myristoylated pseudosubstrate peptide inhibitor (Myr-SIYRRGARRWRKL-OH), which consists of pseudosubstrate peptide sequence derived from PKCz/l and is a cell-permeable, reversible, substrate competitive inhibitor (26). PKCd (V1-1; Myr-SFNSYELGSL-OH) and PKCε (Myr-EAVSLKPT-OH) inhibitors are cell permeable myristoylated translocation peptides inhibitors of PKCd and PKCε (27,28).…”

CpG DNA Activates Survival in Murine Macrophages through TLR9 and the Phosphatidylinositol 3-Kinase-Akt Pathway

“…The pathways leading to MEK-ERK activation by LPS and CSF-1 may be distinct. The involvement of the upstream Ras-Raf components in MEK-ERK activation by LPS is controversial (15,(21)(22)(23)(24), whereas the alternative MAP-3-kinase, Cot/ Tpl-2 was required for LPS-induced MEK-ERK activation (25,26). The pathways leading to PI3K-Akt activation by LPS and CpG DNA still remain poorly defined.…”