Ubiquitination of RIPK1 suppresses programmed cell death by regulating RIPK1 kinase activation during embryogenesis

Xixi Zhang,Ming Li,Wenjuan Pu,Hao Ying,Haikun Wang,Dali Li,Hui Wang,Xiaoming Li,Chengxian Xu,Haiwei Zhang,Qiurong Ding,Haibing Zhang,Xiaoxia Wu,Bin Zhou

doi:10.1038/s41467-019-11839-w

Abstract

The ubiquitination status of RIPK1 is considered to be critical for cell fate determination. However, the in vivo role for RIPK1 ubiquitination remains undefined. Here we show that mice expressing RIPK1K376R which is defective in RIPK1 ubiquitination die during embryogenesis. This lethality is fully rescued by concomitant deletion of Fadd and Ripk3 or Mlkl. Mechanistically, cells expressing RIPK1K376R are more susceptible to TNF-α induced apoptosis and necroptosis with more complex II formation and increased RIPK1 activation, which is consistent with the observation that Ripk1K376R/K376R lethality is effectively prevented by treatment of RIPK1 kinase inhibitor and is rescued by deletion of Tnfr1. However, Tnfr1−/−Ripk1K376R/K376R mice display systemic inflammation and die within 2 weeks. Significantly, this lethal inflammation is rescued by deletion of Ripk3. Taken together, these findings reveal a critical role of Lys376-mediated ubiquitination of RIPK1 in suppressing RIPK1 kinase activity–dependent lethal pathways during embryogenesis and RIPK3-dependent inflammation postnatally.

Highlights

The ubiquitination status of RIPK1 is considered to be critical for cell fate determination
Immunoblot analysis showed activated caspase-3 and the cleavage of PARP, as well as aggregations of RIPK1 and RIPK3 were clearly detected in body tissues of mutant embryos, suggesting that activation of apoptosis and necroptosis contributes to the cell death in mutant embryos (Fig. 1f)
We show that the disruption of RIPK1 ubiquitination in vivo by a point mutation at K376 causes embryonic lethality at E12.5, which is completely different from the observation in Ripk[1] deficient mice

Summary

Introduction

The ubiquitination status of RIPK1 is considered to be critical for cell fate determination. Ablation of Ripk[1] prevents early embryonic lethality induced by Caspase-8 or Fadd, these mice succumb to early postnatal lethality similar to Ripk[1] deficient mice[21,22,25] Another striking study showed that mice with homozygous Ripk3D161N died at E10.5 but were completely rescued by co-deletion of Ripk[126]. A recent work has shown that DD-mediated RIPK1 dimerization is crucial for RIPK1 activation, and mice with mutant DD are resistant to RIPK1-dependent apoptosis and necroptosis[37] These genetic studies have demonstrated that RIPK1 is involved in mediating apoptosis, necroptosis and cell survival, the mechanism by which RIPK1 regulates these pathways in vivo remains unclear

Methods

Results

Conclusion