Abstract
Necroptosis, an inflammatory form of cell death, is initiated by the activation of receptor-interacting protein kinase 3 (RIPK3), which depends on its interaction with RIPK1. Although catalytically inactive, the RIPK3 mutant D161N still stimulates RIPK1-dependent apoptosis and embryonic lethality in RIPK3 D161N homozygous mice. Whereas the absence of RIPK1 rescues RIPK3 D161N homozygous mice, we report that the absence of RIPK1 leads to embryonic lethality in RIPK3 D161N heterozygous mice. This suggested that the kinase domain of RIPK3 had a noncatalytic function that was enhanced by a conformation induced by the D161N mutation. We found that the RIPK3 kinase domain homodimerized through a surface that is structurally similar to that of the RAF family members. Mutation of residues at the dimer interface impaired dimerization and necroptosis. Kinase domain dimerization stimulated the activation of RIPK3 through cis-autophosphorylation. This noncatalytic, allosteric activity was enhanced by certain kinase-deficient mutants of RIPK3, including D161N. Furthermore, apoptosis induced by certain RIPK3 inhibitors was also dependent on the kinase dimerization interface. Our studies reveal that the RIPK3 kinase domain exhibits catalytically independent function that is important for both RIPK3-dependent necroptosis and apoptosis.