title: Stress-induced phosphorylation of PACT reduces its interaction with TRBP and leads to PKR activation.
abstract: PACT is a stress-modulated activator of interferon (IFN)-induced double-stranded (ds) RNA-activated protein kinase (PKR) and is an important regulator of PKR-dependent signaling pathways. Stress-induced phosphorylation of PACT is essential for PACT's association with PKR leading to PKR activation. PKR activation by PACT leads to phosphorylation of translation initiation factor eIF2α, inhibition of protein synthesis, and apoptosis. In addition to positive regulation by PACT, PKR activity in cells is also negatively regulated by TRBP. In this study, we demonstrate for the first time that stress-induced phosphorylation at serine 287 significantly increases PACT's ability to activate PKR by weakening PACT's interaction with TRBP. A non-phosphorylatable alanine substitution mutant at this position causes enhanced interaction of PACT with TRBP and leads to a loss of PKR activation. Furthermore, TRBP overexpression in cells is unable to block apoptosis induced by a phospho-mimetic, constitutively active PACT mutant. These results demonstrate for the first time that stress-induced PACT phosphorylation functions to free PACT from the inhibitory interaction with TRBP and also to enhance its interaction with PKR.
authors.affiliation: Department of Biological Sciences, Developmental Biology and Anatomy, School of Medicine, University of South Carolina, Columbia, SC 29208, USA.
authors.forename: Madhurima
authors.initials: M
authors.lastname: Singh
authors.forename: David
authors.initials: D
authors.lastname: Castillo
authors.forename: Chandrashekhar V
authors.initials: CV
authors.lastname: Patel
authors.forename: Rekha C
authors.initials: RC
authors.lastname: Patel
mesh_terms: D001483:Base Sequence; D017931:DNA Primers; D004789:Enzyme Activation; D015497:HIV-1 / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D010766:Phosphorylation; D011485:Protein Binding; D016601:RNA-Binding Proteins / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D014764:Viral Proteins / Q000378:metabolism*; D019892:eIF-2 Kinase / Q000378:metabolism*
chemical_list: D017931:DNA Primers; C104647:PRKRA protein, human; D016601:RNA-Binding Proteins; D014764:Viral Proteins; D019892:eIF-2 Kinase