Abstract
Necroptosis is a form of regulated necrotic cell death that promotes inflammation. In cells undergoing necroptosis, activated RIPK1 kinase mediates the formation of RIPK1/RIPK3/MLKL complex to promote MLKL oligomerization and execution of necroptosis. RIPK1 kinase activity also promotes cell-autonomous activation of proinflammatory cytokine production in necroptosis. However, the signaling pathways downstream of RIPK1 kinase in necroptosis and how RIPK1 kinase activation controls inflammatory response induced by necroptosis are still largely unknown. Here, we quantitatively measured the temporal dynamics of over 7000 confident phosphorylation-sites during necroptosis using mass spectrometry. Our study defined a RIPK1-dependent phosphorylation pattern in late necroptosis that is associated with a proinflammatory component marked by p-S473 TRIM28. We show that the activation of p38 MAPK mediated by oligomerized MLKL promotes the phosphorylation of S473 TRIM28, which in turn mediates inflammation during late necroptosis. Taken together, our study illustrates a mechanism by which p38 MAPK may be activated by oligomerized MLKL to promote inflammation in necroptosis.
Highlights
Necroptosis is a form of regulated necrosis that can be activated in apoptosis-deficient conditions
We present a quantitative phosphoproteomic study that reveals the dynamic time-dependent changes of RIPK1 kinase-dependent phosphoproteome during necroptosis induced by TNFα in FADDdeficient Jurkat cells
Our study reveals a set of RIPK1-dependent phosphorylation events in late necroptosis
Summary
Necroptosis is a form of regulated necrosis that can be activated in apoptosis-deficient conditions. RIPK1 kinase activity is critical for regulating necroptosis and cell-autonomous cytokine production during necroptosis [1, 2]. Necrostatin-1s (Nec-1s), the first small molecule inhibitor of RIPK1 kinase developed, is an effective inhibitor of necroptosis and inflammation [1, 3, 4]. Necroptosis and RIPK1 kinase-dependent inflammation have been implicated in pathogenesis of various human diseases, including inflammatory bowel disease, liver cancer, and neurodegenerative diseases [5]. Inhibition of RIPK1 using Nec-1s and other small molecule inhibitors of RIPK1 has shown efficacy in various animal models of human diseases [3, 4, 9,10,11,12]. RIPK1 inhibitors have been advanced into human clinical studies for the treatment of human inflammatory and degenerative diseases including inflammatory bowel diseases, rheumatoid arthritis, psoriasis, ALS and AD [5]
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