IFN-γ Induces gp91phox Expression in Human Monocytes via Protein Kinase C-Dependent Phosphorylation of PU.1

Abstract

We previously reported that the stimulation of human blood monocytes with IFN-γ induces the binding of PU.1 to the gp91phox promoter and the consequent expression of gp91phox. In this study, we show that the effect of IFN-γ is reproduced by the serine phosphatase inhibitor, okadaic acid, and this suggests that serine kinases could be involved in gp91phox expression. We also show that IFN-γ induces the serine/threonine phosphorylation of PU.1 in cultured monocytes. This phosphorylation, as well as the IFN-γ-induced PU.1 binding and gp91phox protein synthesis, is slightly affected by the casein kinase II inhibitor, daidzein, but is abrogated by the protein kinase C (PKC) -α and -β inhibitor, Go6976, and by synthetic peptides with sequences based on the endogenous pseudosubstrate region of the classical PKC α and β isoforms. In contrast, peptides reproducing the pseudosubstrate region of PKC ε were without effect. Moreover, we found that the treatment of monocytes with IFN-γ induces the nuclear translocation and the activation of PKC α and βI, but not of PKC βII, and that the IFN-γ-induced phosphorylation of PU.1 was greatly reduced by LY333531, a selective inhibitor of PKC β isoforms. Finally, nuclear run-on assays demonstrated that while the PKC inhibitors, Go6976 and LY333531, decrease the IFN-γ-induced gp91phox transcription, the serine phosphatase inhibitor, okadaic acid, enhances the gp91phox gene transcription. Our results indicate that in cultured monocytes, IFN-γ induces the binding of PU.1 to the gp91phox promoter and the expression of gp91phox by phosphorylation of PU.1 via activation of PKC α and/or βI.

Glucose, palmitate and pro-inflammatory cytokines modulate production and activity of a phagocyte-like NADPH oxidase in rat pancreatic islets and a clonal beta cell line

“…It has been reported that a major source of ROS is the mitochondrion, in which increased availability of substrates causes enhanced oxidative activity and production of oxygen derivatives [12,13]. However, we speculate that a major source of superoxide is the NADPH oxidase system, which may be upregulated in the presence of saturated NEFA, or indeed of proinflammatory cytokines, which are known to increase production of components of the NADPH oxidase system in the phagocytic cells of the immune system [14,15].…”

Mouse neutrophils lacking lamin B-receptor expression exhibit aberrant development and lack critical functional responses

“…Several recent studies of other inner nuclear envelope proteins lend support to this notion; the Xenopus homolog of mammalian MAN1, XMAN1, was shown to block transcriptional regulation by bone morphogenetic protein, and LAP2β repressed the transcriptional activity of several transcription factors, including E2F family members p53 and NF-kB [38][39][40]. Multiple transcriptional regulators have been identified to positively regulate gp91 phox expression, including NF-kB, PU.1, C/EBPε and HoxA9 [41][42][43][44]. It is therefore conceivable that the loss of LBR expression specifically affects the ability of one or more of these factors to properly initiate gp91 phox gene activation.…”

Mechanistic and Therapeutic Studies Related to Anemia of Chronic Disease and Inflammation

“…PKCβ has been shown to undergo recruitment to mitochondrial aconitase [114]; c) PKCα drives macrophage differentiation in a manner similar to PU.1 [85,117]; and d) PKCβ has previously been implicated in direct regulation of PU.1 [75].…”

“…Based on prior implication of PKCβ in the regulation of PU.1 and in the interaction with mitochondrial aconitase [75,114], we predict that PKCβ knockout mice will show in their response to the anemic challenges: diminished erythroid induction of PU.1, diminished impairment of erythropoiesis, and diminished responsiveness to isocitrate treatment.…”

“…Furthermore, shRNA knockdown of mitochondrial aconitase impairs primary erythroid differentiation [28], while knockout of cytosolic aconitase has no effect on steady state or stress erythropoiesis in mice [27]. Previously, we developed a model in which mitochondrial aconitase functions to integrate information on iron availability, [75,76,115]. Furthermore, PKC signaling has been shown to alter hematopoietic cell fate in a manner similar to PU.1 induction [83,85,116].…”