PI3K mediates tumor necrosis factor induced-necroptosis through initiating RIP1-RIP3-MLKL signaling pathway activation.

Necroptosis is a recently identified programmed cell death, which is initiated by receptor-interacting serine/threonine-protein kinase 1 (RIP1), RIP3 and mixed-lineage kinase domain-like protein (MLKL). It has been reported that necroptosis induced by tumor necrosis factor (TNF) was inhibited by the inhibitor of phosphatidylinositol-3-kinase (PI3K) and its substrate protein AKT, indicating that PI3K-AKT signaling pathway was involved in mediating TNF-induced necroptosis, whereas it is unclear how PI3K initiates necroptosis. In this study, we found that TNF-induced necroptosis was inhibited by chemical inhibition or genetic deletion of PI3K. Moreover, knockdown of p110α, the catalytic subunit of PI3K, significantly suppressed the phosphorylation of PI3K substrate protein AKT, and TNF-induced necroptosis was blocked by AKT inhibitors. Furthermore, we found that p110α knockdown also suppressed the phosphorylation and oligomerization of RIP1, RIP3 and MLKL in response to TNF stimulation. In addition to the critical role in mediating TNF-induced necrosome formation, p110α was also essential for the spontaneous phosphorylation of RIP1 and RIP3. Finally, we found that p110α bound to RIP3, but not RIP1, to form protein complex in the process of TNF-induced necroptosis, and mediated TNF-induced necroptosis in the absence of RIP1. Our results demonstrate that PI3K is essential for TNF-induced necroptosis, which may act as the partner of RIP3 to initiate the activation of RIP1-RIP3-MLKL signal pathway and the subsequent necroptosis.