TNFa and IKKb-mediated TANK/I-TRAF phosphorylation: implications for interaction with NEMO/IKKg and NF-kB activation

in Biochemical Journal (2006), 394

Pro-inflammatory cytokines trigger signalling cascades leading to NF-kappaB (nuclear factor-kappaB)-dependent gene expression through IKK [IkappaB (inhibitory kappaB) kinase]-dependent phosphorylation and ... [more ▼]

Pro-inflammatory cytokines trigger signalling cascades leading to NF-kappaB (nuclear factor-kappaB)-dependent gene expression through IKK [IkappaB (inhibitory kappaB) kinase]-dependent phosphorylation and subsequent degradation of the IkappaB proteins and via induced phosphorylation of p65. These signalling pathways rely on sequentially activated kinases which are assembled by essential and non-enzymatic scaffold proteins into functional complexes. Here, we show that the pro-inflammatory cytokine TNFalpha (tumour necrosis factor alpha) promotes TANK [TRAF (TNF receptor-associated factor) family member associated NF-kappaB activator] recruitment to the IKK complex via a newly characterized C-terminal zinc finger. Moreover, we show that TANK is phosphorylated by IKKbeta upon TNFalpha stimulation and that this modification negatively regulates TANK binding to NEMO (NF-kappaB essential modulator). Interestingly, reduced TANK expression by RNA interference attenuates TNFalpha-mediated induction of a subset of NF-kappaB target genes through decreased p65 transactivation potential. Therefore the scaffold protein TANK is required for the cellular response to TNFalpha by connecting upstream signalling molecules to the IKKs and p65, and its subsequent IKKbeta-mediated phosphorylation may be a mechanism to terminate the TANK-dependent wave of NF-kappaB activation. [less ▲]

Association of the adaptor TANK with the IκB kinase (IKK) regulator NEMO connects IKK complexes with IKKε and TBK1 kinases

in Journal of Biological Chemistry (2002), 277(40), 37029-37036

Canonical activation of NF-kappaB is mediated via phosphorylation of the inhibitory IkappaB proteins by the IkappaB kinase complex (IKK). IKK is composed of a heterodimer of the catalytic IKKalpha and ... [more ▼]

Canonical activation of NF-kappaB is mediated via phosphorylation of the inhibitory IkappaB proteins by the IkappaB kinase complex (IKK). IKK is composed of a heterodimer of the catalytic IKKalpha and IKKbeta subunits and a presumed regulatory protein termed NEMO (NF-kappaB essential modulator) or IKKgamma. NEMO/IKKgamma is indispensable for activation of the IKKs in response to many signals, but its mechanism of action remains unclear. Here we identify TANK (TRAF family member-associated NF-kappaB activator) as a NEMO/IKKgamma-interacting protein via yeast two-hybrid analyses. This interaction is confirmed in mammalian cells, and the domains required are mapped. TANK was previously shown to assist NF-kappaB activation in a complex with TANK-binding kinase 1 (TBK1) or IKKepsilon, two kinases distantly related to IKKalpha/beta, but the underlying mechanisms remained unknown. Here we show that TBK1 and IKKepsilon synergize with TANK to promote interaction with the IKKs. The TANK binding domain within NEMO/IKKgamma is required for proper functioning of this IKK subunit. These results indicate that TANK can synergize with IKKepsilon or TBK1 to link them to IKK complexes, where the two kinases may modulate aspects of NF-kappaB activation. [less ▲]