Binding of TNFα to TNF receptor 1 (TNFR1) induces the sequential formation of several signaling complexes, namely complex I and complex IIa/b (Walczak H 2011; Yuan J et al. 2019). These complexes support either cell survival (complex I) or cell death (complex II). The dynamic assembly of these complexes is tightly regulated by proteolysis, ubiquitination and phosphorylation of receptor-interacting serine/threonine protein kinase 1 (RIPK1) and other components of the TNFα signaling pathway (reviewed in Yuan J et al. 2019; Varfolomeev E & Vucic D 2022). RIPK1 functions as a key regulator of both cell survival and cell death (reviewed in Ju E et al. 2022). Enzymatically inactive polyubiquitin-bound RIPK1 serves as a scaffold in the TNFR1 signaling complex (complex I) to recruit TAK1 kinase complex and I-kappa-B kinase (IKK) complex contributing to activation of mitogen-activated protein kinase (MAPK) and NF-kappa-B signaling pathways, which regulate expression of pro-survival and inflammatory genes. Deubiquitination of RIPK1 leads to the activation of RIPK1 kinase activity, which promotes autophosphorylation of RIPK1 and RIPK1 kinase-dependent cell death (reviewed in Ju E et al. 2022). The kinase activity of RIPK1 is regulated by phosphorylation (Lafont E et al. 2018; Dondelinger Y et al. 2019; reviewed in Delanghe T et al. 2020; Ju E et al. 2022). The components of I-kappa-B kinase complex, namely inhibitor of nuclear factor kappa B subunit α (IKKα or CHUK) and subunit β (IKKβ or IKBKB), can directly phosphorylate RIPK1 within the membrane-bound TNFR1 signaling complex (complex I) (Dondelinger Y et al. 2015, 2019). IKK phosphorylates S25 in the kinase domain of RIPK1 to block ATP binding, preventing RIPK1 kinase activation and cell death (Dondelinger Y et al. 2019). Expression of phospho-mimetic S25D RIPK1 mutant prevented TNF-induced cell death in RIPK1-null human Jurkat cells and mouse embryonic fibroblasts (MEF). Autophosphorylation of RIPK1 on S166 and activation of RIPK1 kinase were inhibited in mouse bone marrow-derived macrophages (BMDMs) and MEFs isolated from mice carrying S25D RIPK1 mutant (Dondelinger Y et al. 2019; Puylaert P et al. 2022). Inhibition of IKK activity by Yersinia infection or by SHARPIN gene mutations results in defective phosphorylation of RIPK1 at S25 by IKKα/β (Dondelinger Y et al. 2019). Restoring S25 phosphorylation with S25D RIPK1 prevented TNF-induced RIPK1 kinase-dependent cell death in Yersinia-infected BMDMs. Further, mimicking S25 phosphorylation impaired host immune response to Yersinia infection in vivo in bone marrow chimeric mice, suggesting that RIPK1 kinase-dependent cell death contributes to immune defense in response to pathogens. Similarly, S25D RIPK1 suppressed TNF-induced cell death in mouse dermal fibroblasts (MDFs) isolated from SHARPIN-deficient mice. These data suggest that IKKα/β-mediated phosphorylation of RIPK1 at S25 downregulates RIPK1 auto-phosphorylation downstream of TNFR1, thereby preventing TNFα-induced RIPK1 kinase-dependent cell death (Dondelinger Y et al. 2019).

This Reactome event describes IKK mediated phosphorylation of RIPK1 at S25 within the TNFR1 signaling complex.