Phospholipase C-δ(1) regulates interleukin-1β and tumor necrosis factor-α mRNA expression.

Phospholipase C-δ(1) (PLCδ(1)) is a widely expressed highly active PLC isoform, modulated by Ca(2+) that appears to operate downstream from receptor signaling and has been linked to regulation of cytokine production. Here we investigated whether PLCδ(1) modulated expression of the pro-inflammatory cytokines interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in rat C6 glioma cells. Expression of PLCδ(1) was specifically suppressed by small interfering RNA (siRNA) and the effects on cytokine mRNA expression, stimulated by the Toll-like receptor (TLR) agonist, lipopolysaccharide (LPS), were examined. Real-time polymerase chain reaction (RT-PCR) results showed that PLCδ(1) knockdown enhanced expression IL-1β and tumor necrosis factor-α (TNF-α) mRNA by at least 100 fold after 4 h of LPS stimulation compared to control siRNA treatment. PLCδ(1) knock down caused persistently high Nfκb levels at 4 h of LPS stimulation compared to control siRNA-treated cells. PLCδ(1) knockdown was also associated with elevated nuclear levels of c-Jun after 30 min of LPS stimulation, but did not affect LPS-stimulated p38 or p42/44 MAPK phosphorylation, normally associated with TLR activation of cytokine gene expression; rather, enhanced protein kinase C phosphorylation of cellular proteins was observed in the absence of LPS stimulation. An inhibitor of bisindolylmaleimide II (BIM), reversed phosphorylation, prevented elevation of nuclear c-Jun levels, and inhibited LPS-induced increases of IL-1β and TNF-α mRNA's induced by PLCδ(1) knockdown. Our results show that loss of PLCδ(1) enhances signaling and up-modulates pro-inflammatory cytokine gene transcription in concert with the TLR-stimulated p38MAPK/Nfκb pathway. Our findings are consistent with the idea that PLCδ(1) is a suppressor of activity.

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