Constitutive Association of TGF-β–Activated Kinase 1 with the IκB Kinase Complex in the Nucleus and Cytoplasm of Human Neutrophils and Its Impact on Downstream Processes

Abstract

Neutrophils influence innate and adaptative immunity by generating numerous mediators whose regulation largely depends on the IκB kinase (IKK)/IκB/NF-κB signaling cascade. A singular feature of neutrophils is that they express several components of this pathway (namely, NF-κB/Rel proteins and IκB-α) in both the nucleus and cytoplasm. We recently reported that the IKK complex of neutrophils is similarly expressed and activated in both cellular compartments. However, the upstream IKK kinase has not yet been identified. In this study, we report that neutrophils express the mitogen-activated protein 3 kinase, TGF-β–activated kinase 1 (TAK1), as well as its associated partners, TAK1-binding protein (TAB) 1, TAB2, and TAB4, in both the cytoplasm and nucleus. Following cell stimulation by TNF-α or LPS, TAK1 becomes rapidly and transiently activated. Blocking TAK1 kinase activity with a highly selective inhibitor (5z-7-oxozeaenol) attenuated the phosphorylation of nuclear and cytoplasmic IKKα/β, IκB-α, and RelA, and also impaired IκB-α degradation and NF-κB DNA binding in activated neutrophils. Moreover, TAK1 was found to be involved in the activation of p38 MAPK and ERK, which also influence cytokine generation in neutrophils. As a result, inflammatory cytokine expression and release were profoundly impaired following TAK1 inhibition. Similarly, the delayed apoptosis observed in response to LPS or TNF-α was reversed by TAK1 inhibition. By contrast, IKKγ phosphorylation and STAT1 activation were unaffected by TAK1 inhibition. Our data establish the central role of TAK1 in controlling nuclear and cytoplasmic signaling cascades in primary neutrophils, making it a promising target for therapeutic intervention in view of the foremost role of neutrophils in several chronic inflammatory conditions.