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.

坏死性坏死病是一种细胞死亡的炎症形式，它是由与受体相互作用的蛋白激酶3（RIPK3）激活而引发的，这取决于其与RIPK1的相互作用。尽管没有催化活性，但RIPK3突变体D161N在RIPK3 D161N纯合小鼠中仍刺激RIPK1依赖性凋亡和胚胎致死率。鉴于RIPK1的缺失拯救了RIPK3 D161N纯合子小鼠，我们报道了RIPK1的缺失导致RIPK3 D161N杂合子小鼠的胚胎致死率。这表明RIPK3的激酶结构域具有非催化功能，该功能被D161N突变诱导的构象增强。我们发现，RIPK3激酶结构域通过与RAF家族成员结构相似的表面均二聚化。二聚体界面处残基的突变削弱了二聚化和坏死性坏死。激酶域二聚化通过顺式自磷酸化刺激了RIPK3的活化。这种非催化的变构活性被RIPK3的某些激酶缺陷型突变体（包括D161N）增强。此外，某些RIPK3抑制剂诱导的细胞凋亡也依赖于激酶二聚化界面。我们的研究表明，RIPK3激酶结构域显示出催化独立功能，这对依赖RIPK3的坏死性细胞凋亡和细胞凋亡都很重要。