NF-κB dependent expression of A20 controls IKK repression of RIPK1 dependent cell death in activated T cells

The Inhibitor of Kappa B Kinase (IKK) complex is a critical regulator of canonical NF-κB activation. More recently, RIPK1 has also been identified as a phosphorylation target of the IKK complex, resulting in repression of extrinsic cell death pathways. Our previous work shows that normal thymocyte development is exclusively reliant on repression of TNF triggered cell death pathways by IKK, and that NF-κB signalling is in fact redundant for development. The role of IKK signalling in activated T cells is unclear. To investigate this, we analysed activation of IKK2 deficient TCR transgenic T cells with cognate peptide. While early activation events were normal, proliferation of blasts was impaired. Surprisingly, cell cycle progression in IKK2 KO T cells was unperturbed. Instead, dividing cells were more sensitive to apoptosis triggered by extrinsic cell death pathways, since inhibition of RIPK1 kinase activity almost completely rescued cell survival. Transcriptomic analysis of activated IKK2 deficient T cells revealed defective expression of several NF-κB targets, including Tnfaip3, that encodes A20, a negative regulator of NF-κB in T cells. To test whether A20 expression was required to protect IKK2 deficient T cells from cell death, we generated mice with T cells lacking both A20 and IKK2. Conditional deletion of both Ikk2 and Tnfaip3 in T cells resulted in near complete ablation of peripheral naïve T cells, in contrast to mice lacking one or other gene. Strikingly, this phenotype was completely reversed by inhibition of RIPK1 kinase activity in vivo. Therefore, our data suggests that IKK signalling in T cells protects against RIPK1 dependent death, both by direct phosphorylation of RIPK1 and through NF-κB mediated induction of A20, that we identify for the first time as a modulator of RIPK1 function in T cells. ### Competing Interest Statement The authors have declared no competing interest. * CBM : CARD11 BCL10 MALT1 CIAP : Cellular inhibitor of apoptosis protein IKK : inhibitor of kappa B kinase IKK2i : Inhibitor of kappa B kinase 2 IκB : Inhibitors of kappa B IL-7R : Interleukin-7 receptor Nec1 : Necrostatin-1 RIPK1 : Receptor-interacting protein kinase 1 TAK1 : Transforming growth factor-beta activated kinase 1 TNFR1 : Tumour necrosis factor receptor 1 TRADD : Tumour necrosis factor receptor 1-associated death domain protein TRAF : Tumour necrosis factor receptor-associated factor