Stepwise phosphorylation of p65 promotes NF-κB activation and NK cell responses during target cell recognition

Abstract

NF-κB is a key transcription factor that dictates the outcome of diverse immune responses. How NF-κB is regulated by multiple activating receptors that are engaged during natural killer (NK)-target cell contact remains undefined. Here we show that sole engagement of NKG2D, 2B4 or DNAM-1 is insufficient for NF-κB activation. Rather, cooperation between these receptors is required at the level of Vav1 for synergistic NF-κB activation. Vav1-dependent synergistic signalling requires a separate PI3K-Akt signal, primarily mediated by NKG2D or DNAM-1, for optimal p65 phosphorylation and NF-κB activation. Vav1 controls downstream p65 phosphorylation and NF-κB activation. Synergistic signalling is defective in X-linked lymphoproliferative disease (XLP1) NK cells entailing 2B4 dysfunction and required for p65 phosphorylation by PI3K-Akt signal, suggesting stepwise signalling checkpoint for NF-κB activation. Thus, our study provides a framework explaining how signals from different activating receptors are coordinated to determine specificity and magnitude of NF-κB activation and NK cell responses.

DKK3 attenuates the cytotoxic effect of natural killer cells on CD133+ gastric cancer cells

“…DKK3 is a suppressor in cancer cells, however, we found that DKK3 suppressed the cytotoxicity of NK cells in this study. Erk signaling pathway activation was crucial to cytotoxic degranulation of NK cells . Both in this study and other previous studies, it has been reported that DKK3 inhibited the activation of the Erk pathway in cells .…”

“…Erk signaling pathway activation was crucial to cytotoxic degranulation of NK cells. 27 Both in this study and other previous studies, it has been reported that DKK3 inhibited the activation of the Erk pathway in cells. 28,29 In addition, we found that DKK3 could bind NKG2D and the DKK3-NKG2D complex is difficult to form immunological synapses (IS).…”

Molecular checkpoints controlling natural killer cell activation and their modulation for cancer immunotherapy

“…Using an activation model that combines 2B4 with NKG2D or DNAM-1, we found that SLP-76-Vav1 and NF-κB p65 function as common checkpoints for multiple activation pathways in NK cells (Figure 1). 40, 41 In addition, we recently provided evidence demonstrating E3 ubiquitin ligase c-Cbl and glycogen synthase kinase (GSK)-3β as negative regulators of NK cell activation triggered by multiple activating receptors (Figure 1). 25, 42 Because cancer cells express various and heterogeneous ligands for NK activating receptors, it would be desirable to target and modulate signaling molecules that are common to multiple activating receptors for NK cell activation.…”

“…Rather, their synergistic combination is required for effective NF-κB activation and NK cell responses. 41 This combination relays independent and complementary signals that result in the combined phosphorylation of the ‘upstream' SLP-76-Vav1 and unexpectedly, the ‘downstream' NF-κB p65 subunit. Vav1-dependent synergistic signals are required for p65 phosphorylation and NF-κB activation, which is supported by the stepwise regulation of Vav1 and p65 phosphorylation in NK cells.…”

“…In support, Cbl knockdown markedly enhances NF-κB activation and cytokine production by the co-engagement of NKG2D and 2B4, but not by a single receptor. 41 This ‘quantitative' regulation of signaling by c-Cbl may help to prevent the inadvertent activation of NK cells toward normal healthy cells that express limited ligands for the NK cell activating receptor. In this respect, c-Cbl modulation would provide a promising therapeutic strategy to enhance NK cell activation toward cancer cells.…”