Abstract
Ubiquitous protein kinase CK2 is a key regulator of cell migration, proliferation and tumor growth. CK2 is abundant in retinal astrocytes, and its inhibition suppresses retinal neovascularization in a mouse retinopathy model. In human astrocytes, CK2 co-distributes with GFAP-containing intermediate filaments, which implies its association with cytoskeleton. Contrary to astrocytes, CK2 is co-localized in microvascular endothelial cells (HBMVEC) with microtubules and actin stress fibers, but not with vimentin-containing intermediate filaments. Specific CK2 inhibitors (TBB, TBI, TBCA and DMAT) and nine novel CK2 inhibiting compounds (TID43, TID46, Quinolone-7, Quinolone-39, FNH28, FNH62, FNH64, FNH68 and FNH74) were tested at 10–200 μM for their ability to induce morphological alterations in cultured human astrocytes (HAST-40), and HBMVEC (For explanation of the inhibitor names, see “Methods” section). CK2 inhibitors caused dramatic changes in shape of cultured cells with effective inhibitor concentrations between 50 and 100 μM. Attached cells retracted, acquired shortened processes, and eventually rounded up and detached. CK2 inhibitor-induced morphological alterations were completely reversible and were not blocked by caspase inhibition. However, longer treatment or higher inhibitor concentration did cause apoptosis. The speed and potency of the CK2 inhibitors effects on cell shape and adhesion were inversely correlated with serum concentration. Western analyses showed that TBB and TBCA elicited a significant (about twofold) increase in the activation of p38 and ERK1/2 MAP kinases that may be involved in cytoskeleton regulation. This novel early biological cell response to CK2 inhibition may underlie the anti-angiogenic effect of CK2 suppression in the retina.
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Abbreviations
- CK2:
-
Casein kinase II or 2
- ERK:
-
Extracellular signal-regulated kinase
- GFAP:
-
Glial fibrillar acidic protein
- HBMVEC:
-
Human brain microvascular endothelial cells
- MAPK:
-
Mitogen-activated protein kinase
- TBB:
-
4,5,6,7-Tetrabromobenzotriazole
- ZVD-fmk:
-
Z-Val-DL-Asp-fluoromethylketone
References
Meggio F, Pinna LA (2003) One-thousand-and-one substrates of protein kinase CK2? FASEB J 17:349–368
Guerra B, Issinger O-G (2008) Protein kinase CK2 in human disease. Curr Med Chem 15:1870–1886
Trembley JH, Wang G, Unger G, Slaton J, Ahmed K (2009) Protein kinase CK2 in health and disease: CK2: a key player in cancer biology. Cell Mol Life Sci 66:186–1858
Ljubimov AV, Caballero S, Aoki A, Pinna LA, Grant MB, Castellon R (2004) Involvement of protein kinase CK2 in angiogenesis and retinal neovascularization. Invest Ophthalmol Vis Sci 45:4583–4591
Kramerov AA, Saghizadeh M, Pan H, Kabosova A, Montenarh M, Ahmed K, Penn JS, Chan CK, Hinton DR, Grant MB, Ljubimov AV (2006) Expression of protein kinase CK2 in astroglial cells of normal and neovascularized retina. Am J Pathol 168:1722–1736
Kramerov AA, Saghizadeh M, Caballero S, Shaw LC, Li Calzi S, Bretner M, Montenarh M, Pinna LA, Grant MB, Ljubimov AV (2008) Inhibition of protein kinase CK2 suppresses angiogenesis and hematopoietic stem cell recruitment to retinal neovascularization sites. Mol Cell Biochem 316:177–186
Fruttiger M (2002) Development of the mouse retinal vasculature: angiogenesis versus vasculogenesis. Investig Ophthalmol Vis Sci 43:522–527
Canton DA, Litchfield DW (2006) The shape of things to come: an emerging role for protein kinase CK2 in the regulation of cell morphology and the cytoskeleton. Cell Signal 18:267–275
Wei TQ, Tao M (1993) Human erythrocyte casein kinase II: characterization and phosphorylation of membrane cytoskeletal proteins. Arch Biochem Biophys 307:206–216
Greenquist AC, Wyatt JL, Guatelli JC, Shohet SB (1978) The spectrin phosphorylation reaction in human erythrocytes. Prog Clin Biol Res 20:1–24
Singh TJ, Akatsuka A, Blake KR, Huang KP (1983) Phosphorylation of troponin and myosin light chain by cAMP-independent casein kinase-2 from rabbit skeletal muscle. Arch Biochem Biophys 220:615–622
Luise M, Presotto C, Senter L, Betto R, Ceoldo S, Furlan S, Salvatori S, Sabbadini RA, Salviati G (1993) Dystrophin is phosphorylated by endogenous protein kinases. Biochem J 293:243–247
Vorotnikov AV, Shirinsky VP, Gusev NB (1988) Phosphorylation of smooth muscle caldesmon by three protein kinases: implication for domain mapping. FEBS Lett 236:321–324
Smolock EM, Wang T, Nolt JK, Moreland RS (2007) siRNA knock down of casein kinase 2 increases force and cross-bridge cycling rates in vascular smooth muscle. Am J Physiol Cell Physiol 292:C876–C885
Wang D, Jiang DJ (2009) Protein kinase CK2 regulates cytoskeletal reorganization during ionizing radiation-induced senescence of human mesenchymal stem cells. Cancer Res 69:8200–8207
Lim AC, Tiu SY, Li Q, Qi RZ (2004) Direct regulation of microtubule dynamics by protein kinase CK2. J Biol Chem 279:4433–4439
Moreno FJ, Díaz-Nido J, Jiménez JS, Avila J (1999) Distribution of CK2, its substrate MAP1B and phosphatases in neuronal cells. Mol Cell Biochem 191:201–205
Carneiro AC, Fragel-Madeira L, Silva-Neto MA, Linden R (2008) A role for CK2 upon interkinetic nuclear migration in the cell cycle of retinal progenitor cells. Dev Neurobiol 68:620–631
Golub AG, Yakovenko OY, Bdzhola VG, Sapelkin VM, Zien P, Yarmoluk SM (2006) Evaluation of 3-carboxy-4(1H)-quinolones as inhibitors of human protein kinase CK2. J Med Chem 49:6443–6450
Golub AG, Yakovenko OY, Prykhod’ko AO, Lukashov SS, Bdzhola VG, Yarmoluk SM (2008) Evaluation of 4,5,6,7-tetrahalogeno-1H-isoindole-1,3(2H)-diones as inhibitors of human protein kinase CK2. Biochim Biophys Acta 1784:143–149
Goueli SA, Davis AT, Arfman E, Vessella R, Ahmed K (1990) Monoclonal antibodies against nuclear casein kinase NII (PK-N2). Hybridoma 9:609–618
Yde CW, Frogne T, Lykkesfeldt AE, Fichtner I, Issinger OG, Stenvang J (2007) Induction of cell death in antiestrogen resistant human breast cancer cells by the protein kinase CK2 inhibitor DMAT. Cancer Lett 256:229–237
Pagano MA, Poletto G, Di Maira G, Cozza G, Ruzzene M, Sarno S, Bain J, Elliott M, Moro S, Zagotto G, Meggio F, Pinna LA (2007) Tetrabromocinnamic acid (TBCA) and related compounds represent a new class of specific protein kinase CK2 inhibitors. Chembiochem 8:129–139
Sarno S, Reddy H, Meggio F, Ruzzene M, Davies SP, Donella-Deana A, Shugar D, Pinna LA (2001) Selectivity of 4,5,6,7-tetrabromobenzotriazole, an ATP site-directed inhibitor of protein kinase CK2 (‘casein kinase-2’). FEBS Lett 496:44–48
Di Maira G, Salvi M, Arrigoni G, Marin O, Sarno S, Brustolon F, Pinna LA, Ruzzene M (2005) Protein kinase CK2 phosphorylates and upregulates Akt/PKB. Cell Death Differ 12:668–677
Norris JL, Williams KP, Janzen WP, Hodge CN, Blackwell LJ, Popa-Burke IG (2005) Selectivity of SB203580, SB202190 and other commonly used p38 inhibitors: profiling against a multi-enzyme panel. Lett Drug Des Discov 2:516–521
Jalink K, Eichholtz T, Postma FR, van Corven EJ, Moolenaar WH (1993) Lysophosphatidic acid induces neuronal shape changes via a novel, receptor-mediated signaling pathway: similarity to thrombin action. Cell Growth Differ 4:247–255
Suidan HS, Nobes CD, Hall A, Monard D (1997) Astrocyte spreading in response to thrombin and lysophosphatidic acid is dependent on the Rho GTPase. Glia 21:244–252
Sanchez-Ponce D, Muñoz A, Garrido JJ (2011) Casein kinase 2 and microtubules control axon initial segment formation. Mol Cell Neurosci. doi:10.1016/j.mcn.2010.09.005
Kragh CL, Lund LB, Febbraro F, Hansen HD, Gai WP, El-Agnaf O, Richter-Landsberg C, Jensen PH (2009) α-Synuclein aggregation and Ser-129 phosphorylation-dependent cell death in oligodendroglial cells. J Biol Chem 284:10211–10222
Leung YM, Kwan TK, Kwan CY, Daniel EE (2002) Calyculin A-induced endothelial cell shape changes are independent of [Ca2+](i) elevation and may involve actin polymerization. Biochim Biophys Acta 1589:93–103
Parizi M, Howard EW, Tomasek JJ (2000) Regulation of LPA-promoted myofibroblast contraction: role of Rho, myosin light chain kinase, and myosin light chain phosphatase. Exp Cell Res 254:210–220
Li X, Zhou L, Gorodeski GI (2006) Estrogen regulates epithelial cell deformability by modulation of cortical actomyosin through phosphorylation of nonmuscle myosin heavy-chain II-B filaments. Endocrinology 147:5236–5248
Goedert M, Hasegawa M, Jakes R, Lawler S, Cuenda A, Cohen P (1997) Phosphorylation of microtubule-associated protein tau by stress-activated protein kinases. FEBS Lett 409:57–62
Woo MS, Ohta Y, Rabinovitz I, Stossel TP, Blenis J (2004) Ribosomal S6 kinase (RSK) regulates phosphorylation of filamin A on an important regulatory site. Mol Cell Biol 24:3025–3035
Cheng TJ, Lai YK (1998) Identification of mitogen-activated protein kinase-activated protein kinase-2 as a vimentin kinase activated by okadaic acid in 9L rat brain tumor cells. J Cell Biochem 71:169–181
Rouse J, Cohen P, Trigon S, Morange M, Alonso-Llamazares A, Zamanillo D, Hunt T, Nebreda AR (1994) A novel kinase cascade triggered by stress and heat shock that stimulates MAPKAP kinase-2 and phosphorylation of the small heat shock proteins. Cell 78:1027–1037
Singh S, Powell DW, Rane MJ, Millard TH, Trent JO, Pierce WM, Klein JB, Machesky LM, McLeish KR (2003) Identification of the p16-Arc subunit of the Arp 2/3 complex as a substrate of MAPK-activated protein kinase 2 by proteomic analysis. J Biol Chem 278:36410–36417
Sayed M, Kim SO, Salh BS, Issinger OG, Pelech SL (2000) Stress-induced activation of protein kinase CK2 by direct interaction with p38 mitogen-activated protein kinase. J Biol Chem 275:16569–16573
Hildesheim J, Salvador JM, Hollander MC, Fornace AJ Jr (2005) Casein kinase 2- and protein kinase A-regulated adenomatous polyposis coli and beta-catenin cellular localization is dependent on p38 MAPK. J Biol Chem 280:17221–17226
Ji H, Wang J, Nika H, Hawke D, Keezer S, Ge Q, Fang B, Fang X, Fang D, Litchfield DW, Aldape K, Lu Z (2009) EGF-induced ERK activation promotes CK2-mediated disassociation of alpha-Catenin from beta-Catenin and transactivation of beta-Catenin. Mol Cell 36:547–559
Zhang H, Shi X, Hampong M, Blanis L, Pelech S (2001) Stress-induced inhibition of ERK1 and ERK2 by direct interaction with p38 MAP kinase. J Biol Chem 276:6905–6908
McKay MM, Ritt DA, Morrison DK (2009) Signaling dynamics of the KSR1 scaffold complex. Proc Natl Acad Sci USA 106:11022–11027
Ramos JW (2008) The regulation of extracellular signal-regulated kinase (ERK) in mammalian cells. Int J Biochem Cell Biol 40:2707–2719
Ritt DA, Zhou M, Conrads TP, Veenstra TD, Copeland TD, Morrison DK (2007) CK2 is a component of the KSR1 scaffold complex that contributes to Raf kinase activation. Curr Biol 17:179–184
Lebrin F, Bianchini L, Rabilloud T, Chambaz EM, Goldberg Y (1999) CK2α-protein phosphatase 2A molecular complex: possible interaction with the MAP kinase pathway. Mol Cell Biochem 191:207–212
Llorens F, Miró FA, Casañas A, Roher N, Garcia L, Plana M, Gómez N, Itarte E (2004) Unbalanced activation of ERK1/2 and MEK1/2 in apigenin-induced HeLa cell death. Exp Cell Res 299:15–26
Van Dross RT, Hong X, Pelling JC (2005) Inhibition of TPA-induced cyclooxygenase-2 expression by apigenin through downregulation of Akt signal transduction in human keratinocytes. Mol Carcinog 44:83–91
Lin JK, Chen YC, Huang YT, Lin-Shiau SY (1997) Suppression of protein kinase C and nuclear oncogene expression as possible molecular mechanisms of cancer chemoprevention by apigenin and curcumin. J Cell Biochem 28:39–48
Pagano MA, Bain J, Kazimierczuk Z, Sarno S, Ruzzene M, Di Maira G, Elliott M, Orzeszko A, Cozza G, Meggio F, Pinna LA (2008) The selectivity of inhibitors of protein kinase CK2: an update. Biochem J 415:353–365
Acknowledgments
This work was supported by R01 EY13431 (AAK, AVL); M01 RR00425; Winnick Family Foundation (AVL); Department of Surgery, Cedars-Sinai Medical Center (AAK, AVL); OneSight Research Foundation (AVL); Eye Defects Research Foundation (AAK, AVL); grant 0107U004939 from the National Academy of Sciences of Ukraine (AGG, VGB, SMY); NIH grant UO1-CA15062 and V.A. Medical Research Funds (KA); grant from Warsaw University of Technology (MB).
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Kramerov, A.A., Golub, A.G., Bdzhola, V.G. et al. Treatment of cultured human astrocytes and vascular endothelial cells with protein kinase CK2 inhibitors induces early changes in cell shape and cytoskeleton. Mol Cell Biochem 349, 125–137 (2011). https://doi.org/10.1007/s11010-010-0667-3
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DOI: https://doi.org/10.1007/s11010-010-0667-3