Stress-induced Activation of Protein Kinase CK2 by Direct Interaction with p38 Mitogen-activated Protein Kinase

Abstract

Protein kinase CK2 has been implicated in the regulation of a wide range of proteins that are important in cell proliferation and differentiation. Here we demonstrate that the stress signaling agents anisomycin, arsenite, and tumor necrosis factor-␣ stimulate the specific enzyme activity of CK2 in the human cervical carcinoma HeLa cells by up to 8-fold, and this could be blocked by the p38 MAP kinase inhibitor SB203580. We show that p38␣ MAP kinase, in a phosphorylation-dependent manner, can directly interact with the ␣ and ␤ subunits of CK2 to activate the holoenzyme through what appears to be an allosteric mechanism. Furthermore, we demonstrate that anisomycin-and tumor necrosis factor-␣-induced phosphorylation of p53 at Ser-392, which is important for the transcriptional activity of this growth suppressor protein, requires p38 MAP kinase and CK2 activities.The signaling pathways that regulated protein kinase CK2 1 (formerly casein kinase II) have been elusive since the discovery of this highly conserved and ubiquitous protein-serine/ threonine kinase over 45 years ago (1-5). Despite the identification of over 160 putative substrates for CK2, the overriding function of this kinase has been evasive. In most cells, the holoenzyme form of CK2 is a constitutively active heterotetramer of two catalytic ␣ and/or ␣Ј) and two regulatory (␤) subunits, but CK2␤-free pools of CK2␣ also exist. Further increases (2-3-fold) in the phosphotransferase activity of CK2 in cell lysates toward synthetic peptide substrates have been reported following treatment of murine and human fibroblastic cell lines, A431 and 3T3-L1 adipocytes with serum (6), insulin (7-9), insulin-like growth factor-18 (10), epidermal growth factor (7, 11), bombesin (12), and tumor necrosis factor (TNF) (13). CK2 phosphotransferase activity and protein levels are also commonly elevated in solid human tumors and transformed cell lines (2-4, 14, 15). Furthermore, CK2␣ overexpression along with c-Myc induces lymphomas in mice (16). While the increased expression of CK2 is linked with neoplastic transformation, little is known concerning acute regulation of this protein kinase. EXPERIMENTAL PROCEDURESCell Culture and Lysates Preparation-HeLa cells were grown at 37°C on 125-cm 2 flasks in M199 medium (Life Technologies, Inc.) supplemented with 10% fetal calf serum. For cell stimulation, HeLa cells were split at 2 ϫ 10 6 cells/10-cm 2 dish 24 h prior to starvation. Cells were starved with 0.5% fetal calf serum for 15-20 h before exposure to stimuli. Starved cells were incubated with 10 M SB203580 (Calbiochem) for 30 min prior to addition of 10 g/ml anisomycin (Sigma) for 30 min, 50 M arsenite (Sigma) for 15 min, or 20 ng/ml TNF␣ for 15 min. Adhering cells were washed twice with ice-cold phosphate-buffered saline, scraped, then lysed in 50 mM Hepes (pH 7.2), 100 mM NaCl, 1 mM EGTA, and 20 mM sodium fluoride in Buffer A (1 mM sodium orthovanadate, 1% aprotinin, 1 mM phenylmethylsulfonyl fluoride, 1 M pepstatin, 0.5 g/ml leupeptin, 10 g/ml soybean trypsin inhibitor, ...

T-Type Ca2+ Channel Inhibition Induces p53-Dependent Cell Growth Arrest and Apoptosis through Activation of p38-MAPK in Colon Cancer Cells

“…4A). Activated p38-MAPK associates with CK2 to enhance phosphorylation of p53 at Ser392 (31)(32)(33). Therefore, we investigated whether p38-MAPK was involved in the The plot represents data from at least three independent experiments AE SEM.…”

“…In particular, upon UV-induced DNA damage or osmotic shock, p38-MAPK phosphorylates p53 at Ser46 (29) or Ser33 (30), thus stabilizing p53. Moreover, UVinduced DNA damage activates p38-MAPK to activate casein kinase 2 (CK2), which in turn also phosphorylates p53 at Ser392, increasing p53 transcriptional activity (31)(32)(33). Thus, p38-MAPK supports increased p53 levels and activity.…”

Phosphorylation regulates the stability of the regulatory CK2β subunit

“…More detailed investigations of CK2 in yeast and in mammalian cells have further demonstrated that CK2 is required for cell cycle progression with roles at di erent stages in the cell cycle including G1, G1/S and G2/M (Hanna et al, 1995;Pepperkok et al, 1991;Lorenz et al, 1993). A role for CK2 in cellular responses to various stresses including UV, anisomycin, arsenite, heat shock and TNFa has also recently emerged (Sayed et al, 2000;Ghavidel and Schultz, 2001;Keller et al, 2001;Gerber et al, 2000).…”

Mutation at p53 serine 389 does not rescue the embryonic lethality in mdm2 or mdm4 null mice

“…Serine 389 in mouse p53 is phosphorylated by casein kinase II and p38 MAP kinase after UV radiation but not IR (Kapoor and Lozano, 1998;Lu et al, 1998;Sayed et al, 2000;Keller et al, 2001). First, we determined if the mutant p53 S389A protein, expressed by the mouse embryonic fibroblasts (MEFs) from a mouse homozygous for the p53 S389A mutation, could not be phosphorylated at codon 389 after UV radiation.…”

“…The extreme C-terminus of p53 is believed to function as a negative regulatory domain (Hupp and Lane, 1994a, b;Pellegata et al, 1995;Wiederschain et al, 2001). Serine 392 in human p53 (389 in mouse) is present in this region and is known to be modified only by ultraviolet (UV) radiation, but not g-radiation (IR), through phosphorylation by casein kinase II and p38 MAP kinase (Kapoor and Lozano, 1998;Lu et al, 1998;Sayed et al, 2000;Keller et al, 2001). Biologically, this modification is known to stabilize the p53 tetramer and enhance sequence-specific DNA-binding ability of p53 through a conformational change Lane, 1994a, 1995;Sakaguchi et al, 1997).…”