Graphical abstract

Allantopyrone A is a fungal metabolite that uniquely possesses two α,β-unsaturated carbonyl moieties. We recently reported that allantopyrone A inhibited the nuclear factor-κB (NF-κB) signaling pathway induced by tumor necrosis factor (TNF)-α in human lung carcinoma A549 cells. In the present study, the mechanism by which allantopyrone A inhibits the TNF-α-induced signaling pathway was investigated in more detail. Allantopyrone A blocked extensive modifications to receptor-interacting protein 1 (RIP1) in the TNF receptor 1 (TNF-R1) complex. Allantopyrone A augmented the high-MW bands of TNF-R1, TNF receptor-associated factor 2, RIP1, the NF-κB subunit RelA and inhibitor of NF-κB kinase β in A549 cells, suggesting that it binds to and promotes the crosslinking of these proteins. The extracellular cysteine-rich domains of TNF-R1 were crosslinked by allantopyrone A more preferentially than its intracellular portion. The present results demonstrate that allantopyrone A interferes with multiple components of the TNF-R1 complex and blocks RIP1 modifications in the TNF-α-induced NF-κB signaling pathway.

Tumor necrosis factor α (TNF-α) induces the activation of transcription factor nuclear factor κB (NF-κB), which upregulates a variety of genes, including the gene encoding intercellular adhesion molecule-1 (ICAM-1). Allantopyrone A, a recently identified α-pyrone metabolite from an endophytic fungus, was found to inhibit the TNF-α-induced expression of ICAM-1 in human lung carcinoma A549 cells. Allantopyrone A also inhibited the TNF-α-induced luciferase expression of an NF-κB-responsive reporter. In the NF-κB signaling pathway, allantopyrone A inhibited the nuclear translocation of NF-κB subunits as well as the phosphorylation and subsequent degradation of the inhibitor of NF-κB (IκB) α proteins. By contrast, allantopyrone A did not directly affect the catalytic activity of active IκB kinase β. These findings indicate that allantopyrone A inhibits the NF-κB signaling pathway at a step upstream of IκBα phosphorylation.