ATR-mediated phosphorylation of RIPK1 inhibits DNA damage-induced necroptosis

Abstract

Necroptosis induced by DNA damage during chemotherapy is a significant and effective treatment strategy for malignant tumors. Ataxia telangiectasia and rad3-related protein (ATR), a key kinase in DNA damage checkpoints, initiates repair by transmitting damage signals to effectors. However, persistent DNA damage may result in cell death. The mechanisms by which ATR regulates necroptosis remain incompletely understood. In this study, we demonstrated that ATR binds to receptor-interacting serine/threonine-protein kinase 1 (RIPK1) and inhibits its activation, thereby suppressing RIPK1-dependent necroptosis triggered by DNA damage. Mechanistically, ATR directly inhibited RIPK1 and downstream necrosome formation through Ser335 phosphorylation following DNA damage, thereby attenuating RIPK1-dependent necroptosis. In the case of the S335A mutation, RIPK1 repression was relieved, leading to enhanced downstream necroptosis. Furthermore, RIPK1 knockout with complementation of wild-type or S335A mutation in ovarian cancer cell lines revealed that ATR phosphorylation of RIPK1 at S335 promoted chemoresistance, while the S335A mutation significantly increased chemosensitivity. This was characterized by heightened necroptosis activation, reduced cell viability, and increased cell death. These findings expand our understanding of the interaction between DNA damage and cell death regulation and may aid in developing therapeutic drugs to enhance DNA damage-induced tumor necroptosis and improve chemosensitivity.