Phosphorylation of IκBα in the C-Terminal PEST Domain by Casein Kinase II Affects Intrinsic Protein Stability

• DOI: 10.1128/mcb.16.4.1401
• Publication Date: 2015-10-09T00:23:17Z
• Authors: Rongtuan Lin; Pierre Beauparlant; Constantin Makris; Sylvain Meloche; John Hiscott
• Keywords: Jurkat cells; Casein kinase 2

The NF-kappaB/Rel transcription factors participate in the activation of immune system regulatory genes and viral early including human immunodeficiency virus type 1 long terminal repeat. proteins are coupled to inhibitory molecules, collectively termed IkappaB, which responsible for cytoplasmic retention NF-kappaB. Cell leads phosphorylation degradation IkappaBalpha, permitting NG-kappaB/Rel translocation nucleus target gene activation. To further characterize signaling events that contribute IkappaBalpha phosphorylation, a kinase activity was isolated from Jurkat T cells specifically interacted with an affinity chromatography step phosphorylated high specificity vitro. By using in-gel assay recombinant as substrate, two forms (43 38 kDa) were identified. Biochemical criteria immunological cross-reactivity identified alpha catalytic subunit casein II (CKII). Deletion mutants delta1 delta4) localized C-terminal PEST domain IkappaBalpha. Point mutation residues T-291, S-283, T-299 dramatically reduced by NIH-3T3 stably expressed wild-type (wtIkappaB), double-point-mutated (T291A, S283A), or triple-point-mutated S283A, T299A) under control tetracycline-responsive promoter generated. Constitutive triple point mutant eliminated vivo, although tumor necrosis factor-inducible unaffected. In cell lines transiently transfected cells, CKII sites resulted protein increased intrinsic stability. Together results demonstrating role N-terminal inducer-mediated these studies indicate important constitutive stability