Key Laboratory of Nutrition and Metabolism, Institute for Nutritional Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai 200031, China.
Department of Plastic and Reconstructive Surgery, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University, Shanghai 200011, China.
Cell Death Differ. 2017 Aug;24(8):1459-1469. doi: 10.1038/cdd.2017.78. Epub 2017 Jun 2.
RIP1 is an essential regulator of TNF-induced signaling complexes mediating NF-κB activation, apoptosis and necroptosis. Loss of Rip1 rescues the embryonic lethality of Fadd or Caspase-8-deficient mice, even though the double knockout mice die shortly after birth like Rip1-deficient mice. Recent studies demonstrated that mice expressing RIP1 kinase-dead mutants developed normally and resisted necroptotic stimuli in vitro and in vivo. However, the impact of RIP1 kinase activity on Fadd embryonic development remains unknown. Here, we engineered two RIP1 kinase inactive mutant mouse lines, a Rip1 mouse line as previously reported and a novel Rip1 mouse line with an altered P-loop in the kinase domain. While RIP1 could not rescue the embryonic lethality of Fadd-deficient mice at E11.5, RIP1 rescued lethality of Fadd mice at E11.5 and FaddRip1 mice eventually died at E16.5 due to excessive death of fetal liver cells and unregulated inflammation. Under necropotosis-inducing conditions, comparing to Rip1 cells, Rip1cells displayed reduced phosphorylation and oligomerization of RIP3 and MLKL, which lead to increased cell viability. Thus, our study provides genetic evidence that different kinase inactive mutations have distinct impacts on the embryogenesis of Fadd-deficient mice, which might attribute to their extents of protection on necroptosis signaling.
RIP1 是 TNF 诱导的信号复合物的必需调节剂，介导 NF-κB 激活、细胞凋亡和坏死性凋亡。Rip1 的缺失挽救了 Fadd 或 Caspase-8 缺陷型小鼠的胚胎致死性，尽管双敲除小鼠在出生后不久就像 Rip1 缺陷型小鼠一样死亡。最近的研究表明，表达 RIP1 激酶失活突变体的小鼠正常发育，并在体外和体内抵抗坏死性凋亡刺激。然而，RIP1 激酶活性对 Fadd 胚胎发育的影响尚不清楚。在这里，我们构建了两条 RIP1 激酶失活突变体的小鼠系，一条是以前报道的 Rip1 小鼠系，另一条是激酶结构域中 P 环改变的新型 Rip1 小鼠系。虽然 RIP1 不能挽救 E11.5 时 Fadd 缺陷型小鼠的胚胎致死性，但 RIP1 挽救了 Fadd 小鼠的致死性，而 FaddRip1 小鼠最终在 E16.5 因胎肝细胞过度死亡和不受调节的炎症而死亡。在诱导坏死性凋亡的条件下，与 Rip1 细胞相比，Rip1 细胞显示出 RIP3 和 MLKL 的磷酸化和寡聚化减少，这导致细胞活力增加。因此，我们的研究提供了遗传证据，表明不同的激酶失活突变对 Fadd 缺陷型小鼠的胚胎发生有不同的影响，这可能归因于它们对坏死性凋亡信号的保护程度。
