Interaction Of Ripk1 And A20 Modulates Mapk Signaling In Murine Acetaminophen Toxicity

Acetaminophen (APAP)-induced liver necrosis is a form of regulated cell death (RCD) in which APAP activates the mitogen-activated protein kinases (MAPKs) and specifically the c-Jun-N-terminal kinase (JNK) pathway, leading to necrotic cell death. Previously, we have shown that receptor interacting protein kinase-1 (RIPK1) knockdown is also protective against APAP RCD upstream of JNK. However, whether the kinase or platform function of RIPK1 is involved in APAP RCD is not known. To answer this question, we used genetic mouse models of targeted hepatocyte RIPK1 knockout (RIPK1(HepCKO)) or kinase dead knock-in (RIPK1(D138N)) and adult hepatocyte specific knockout of the cytoprotective protein A20 (A20(HepCKO)), known to interact with RIPK1, to study its potential involvement in MAPK signaling. We observed no difference in injury between WT and RIPK1D138N mice post APAP. However, RIPK1(HepCKO) was protective. We found that RIPK1(HepCKO) mice had attenuated pJNK activation, while A20 was simultaneously upregulated. Conversely, A20(HepCKO) markedly worsened liver injury from APAP. Mechanistically, we observed a significant upregulation of apoptosis signal-regulating kinase 1 (ASK1) and increased JNK activation in A20(HepCKO) mice compared with littermate controls. We also demonstrated that A20 coimmunoprecipitated (co-IP) with both RIPK1 and ASK1, and that in the presence of RIPK1, there was less A20-ASK1 association than in its absence. We conclude that the kinase-independent platform function of RIPK1 is involved in APAP toxicity. Adult RIPK1(HepCKO) mice are protected against APAP by upregulating A20 and attenuating JNK signaling through ASK1, conversely, A20(HepCKO) worsens injury from APAP.