A study on the essential role of casein kinase gamma 1 and 3 in necroptosis

Abstract

Upon necroptosis activation, receptor interacting serine/threonine kinase<br/> (RIPK) 1 and RIPK3 form a necrosome complex with pseudokinase mixed<br/> lineage kinase-like (MLKL). Although protein phosphorylation is a key<br/> event for the activation of RIPK1 and RIPK3 in response to necroptosis <br/> signal, relatively little is known about other factors that can regulate the<br/> activity of those kinases or necrosome formation. In a gain-of-function<br/> screen with 650 kinases and 120 phosphatases, I identified that casein<br/> kinase 1 gamma (CK1γ) as a crucial regulator of necroptosis. Here I show<br/> that the downregulation of CK1γ1 and CK1γ3, either by a chemical<br/> inhibitor or knockdown in cells, reduced TNFα-induced necroptosis.<br/> Conversely, ectopic expression of CK1γ1 or CK1γ3 exacerbated<br/> necroptosis, but not apoptosis. Like RIPK1 and RIPK3, CK1γ1 was<br/> cleaved at Asp434 by caspase-8 during apoptosis, while it was increased in<br/> response to necroptosis. CK1γ1 and CK1γ3 formed a protein complex with<br/> each other and were recruited into the necrosome harboring RIPK1, RIPK3<br/> and MLKL. Especially, autophosphorylated form of CK1γ3 at Ser344/345<br/> was detected in the necrosome and was required to mediate the necroptosis.<br/> In addition, CK1γ phosphorylates RIPK3 in vitro and a CK1γ-specific<br/> inhibitor Gi reduced the phosphorylation of MLKL at Ser358, as well as <br/> the formation of MLKL oligomers, and rescued mice from TNFα-induced<br/> systemic inflammatory response syndrome (SIRS). Collectively, these data<br/> suggest that CK1γ1 and CK1γ3 are required for promoting cell death<br/> progression by regulating the formation of necrosome through RIPK3.