K63-linked ubiquitination regulates RIPK1 kinase activity to prevent cell death during embryogenesis and inflammation

Area of Science:

• Cell Biology
• Immunology
• Molecular Biology

Background:

• Receptor-interacting protein kinase 1 (RIPK1) is crucial for regulating cell death pathways.
• The precise mechanisms controlling RIPK1 kinase activity and its role in cell death remain incompletely understood.

Purpose of the Study:

• To investigate the role of Lysine 63 (K63)-linked ubiquitination at residue 376 of RIPK1 in regulating its kinase activity.
• To elucidate the impact of impaired K63-linked ubiquitination on RIPK1 in embryonic development and inflammatory responses.

Main Methods:

• Generation of Ripk1^K376R/K376R knock-in mice to impair K63-linked ubiquitination of RIPK1.
• Analysis of embryonic lethality, cell death, and inflammatory markers in knock-in mice and their genetic variants.
• Utilizing TNFα stimulation and genetic deficiencies in TNFR1, RIPK3, and Caspase8 to dissect signaling pathways.

Main Results:

• Ripk1^K376R/K376R mice exhibited early embryonic lethality due to excessive cell death, linked to reduced TAK1 suppression and increased TNFR1 complex II formation.
• Complete prevention of embryonic lethality was observed in Ripk1^K376R/K376R mice lacking RIPK3 and Caspase8.
• Viable Ripk1^K376R/- mice displayed severe systemic inflammation, primarily mediated by RIPK3-dependent signaling, highlighting K63-linked ubiquitination's role in inflammation.

Conclusions:

• K63-linked ubiquitination on RIPK1's Lys376 residue is essential for regulating RIPK1 kinase activity.
• This ubiquitination critically controls cell death programs and prevents embryonic lethality.
• Impaired K63-linked ubiquitination of RIPK1 contributes to inflammatory processes, underscoring its dual role in cell death and immunity.