Cellular and Molecular Life Sciences

, Volume 66, Issue 13, pp 2181–2193

Calcium signaling-induced Smad3 nuclear accumulation induces acetylcholinesterase transcription in apoptotic HeLa cells

Research Article

Abstract

Recently, acetylcholinesterase (AChE) has been studied as an important apoptosis regulator. We previously showed that cellular calcium mobilization upregulated AChE expression by modulating promoter activity and mRNA stability. In this study, we have identified a potential Smad3/4 binding element, TGCCAGACA, located within the −601 to −571 bp fragment of the AChE promoter, as an important calcium response motif. Smad2/3 and Smad4 were shown to bind this element. Overexpression of human Smad3 increased AChE transcription activity in a dose-dependent manner in HeLa cells, whereas dominant-negative Smad3 blocked this activation. Upon A23187 and thapsigargin treatment, nuclear Smad3 accumulation was observed, an effect that was blocked by the intracellular Ca2+ chelator BAPTA–AM. Calcium-induced AChE transcriptional activation was significantly blocked when the nuclear localization signal of Smad3 was destroyed. Taken together, our data suggest Smad3 can regulate AChE transcriptional activation following calcium-induced nuclear accumulation.

Keywords

Smad3 Acetylcholinesterase Cellular calcium perturbation Apoptosis Nuclear accumulation Transcriptional activation 

References

  1. 1.
    Taylor P, Radic Z (1994) The cholinesterases: from genes to proteins. Annu Rev Pharmacol Toxicol 34:281–320PubMedCrossRefGoogle Scholar
  2. 2.
    Zhang XJ, Yang L, Zhao Q, Caen JP, He HY, Jin QH, Guo LH, Alemany M, Zhang LY, Shi YF (2002) Induction of acetylcholinesterase expression during apoptosis in various cell types. Cell Death Differ 9:790–800PubMedCrossRefGoogle Scholar
  3. 3.
    Jiang H, Zhang XJ (2008) Acetylcholinesterase and apoptosis. A novel perspective for an old enzyme. FEBS J 275:612–617PubMedCrossRefGoogle Scholar
  4. 4.
    Jin QH, He HY, Shi YF, Lu H, Zhang XJ (2004) Overexpression of acetylcholinesterase inhibited cell proliferation and promoted apoptosis in NRK cells. Acta Pharmacol Sin 25:1013–1021PubMedGoogle Scholar
  5. 5.
    Yang L, He HY, Zhang XJ (2002) Increased expression of intranuclear AChE involved in apoptosis of SK-N-SH cells. Neurosci Res 42:261–268PubMedCrossRefGoogle Scholar
  6. 6.
    Park SE, Kim ND, Yoo YH (2004) Acetylcholinesterase plays a pivotal role in apoptosome formation. Cancer Res 64:2652–2655PubMedCrossRefGoogle Scholar
  7. 7.
    Toiber D, Berson A, Greenberg D, Melamed-Book N, Diamant S, Soreq H (2008) N-acetylcholinesterase-induced apoptosis in Alzheimer’s disease. PLoS ONE 3:e3108PubMedCrossRefGoogle Scholar
  8. 8.
    Demaurex N, Distelhorst C (2003) Cell biology. Apoptosis—the calcium connection. Science 300:65–67PubMedCrossRefGoogle Scholar
  9. 9.
    Mattson MP, Chan SL (2003) Calcium orchestrates apoptosis. Nat Cell Biol 5:1041–1043PubMedCrossRefGoogle Scholar
  10. 10.
    Virgilio F, Pozzan T (2003) Calcium and apoptosis: facts and hypotheses. Oncogene 22:8619–8627PubMedCrossRefGoogle Scholar
  11. 11.
    Groenendyk J, Lynch J, Michalak M (2004) Calreticulin, Ca2+, and calcineurin—signaling from the endoplasmic reticulum. Mol Cells 17:383–389PubMedGoogle Scholar
  12. 12.
    Luo Z, Fuentes ME, Taylor P (1994) Regulation of acetylcholinesterase mRNA stability by calcium during differentiation from myoblasts to myotubes. J Biol Chem 269:27216–27223PubMedGoogle Scholar
  13. 13.
    Sberna G, Saez-Valero J, Beyreuther K, Masters CL, Small DH (1997) The amyloid beta-protein of Alzheimer’s disease increases acetylcholinesterase expression by increasing intracellular calcium in embryonal carcinoma P19 cells. J Neurochem 69:1177–1184PubMedGoogle Scholar
  14. 14.
    Choi RC, Siow NL, Cheng AW, Ling KK, Tung EK, Simon J, Barnard EA, Tsim KW (2003) ATP acts via P2Y1 receptors to stimulate acetylcholinesterase and acetylcholine receptor expression: transduction and transcription control. J Neurosci 23:4445–4456PubMedGoogle Scholar
  15. 15.
    Jing P, Jin Q, Wu J, Zhang XJ (2008) GSK3beta mediates the induced expression of synaptic acetylcholinesterase during apoptosis. J Neurochem 104:409–419PubMedGoogle Scholar
  16. 16.
    Zhu H, Gao W, Jiang H, Jin QH, Shi YF, Tsim KW, Zhang XJ (2007) Regulation of acetylcholinesterase expression by calcium signaling during calcium ionophore A23187- and thapsigargin-induced apoptosis. Int J Biochem Cell Biol 39:93–108PubMedCrossRefGoogle Scholar
  17. 17.
    Zhu H, Gao W, Jiang H, Wu J, Shi YF, Zhang XJ (2007) Calcineurin mediates acetylcholinesterase expression during calcium ionophore A23187-induced HeLa cell apoptosis. Biochim Biophys Acta 1773:593–602PubMedCrossRefGoogle Scholar
  18. 18.
    Zhu H, Gao W, Shi YF, Zhang XJ (2007) The CCAAT-binding factor CBF/NF-Y regulates the human acetylcholinesterase promoter activity during calcium ionophore A23187-induced cell apoptosis. Biochim Biophys Acta 1770:1475–1482PubMedGoogle Scholar
  19. 19.
    Feng XH, Derynck R (2005) Specificity and versatility in tgf-beta signaling through Smads. Annu Rev Cell Dev Biol 21:659–693PubMedCrossRefGoogle Scholar
  20. 20.
    Shi Y, Massague J (2003) Mechanisms of TGF-beta signaling from cell membrane to the nucleus. Cell 113:685–700PubMedCrossRefGoogle Scholar
  21. 21.
    Shi Y, Wang YF, Jayaraman L, Yang H, Massague J, Pavletich NP (1998) Crystal structure of a Smad MH1 domain bound to DNA: insights on DNA binding in TGF-beta signaling. Cell 94:585–594PubMedCrossRefGoogle Scholar
  22. 22.
    Inman GJ, Nicolas FJ, Hill CS (2002) Nucleocytoplasmic shuttling of Smads 2, 3, and 4 permits sensing of TGF-beta receptor activity. Mol Cell 10:283–294PubMedCrossRefGoogle Scholar
  23. 23.
    Xiao Z, Liu X, Henis YI, Lodish HF (2000) A distinct nuclear localization signal in the N terminus of Smad 3 determines its ligand-induced nuclear translocation. Proc Natl Acad Sci USA 97:7853–7858PubMedCrossRefGoogle Scholar
  24. 24.
    Schmierer B, Hill CS (2007) TGFbeta-SMAD signal transduction: molecular specificity and functional flexibility. Nat Rev Mol Cell Biol 8:970–982PubMedCrossRefGoogle Scholar
  25. 25.
    Kretzschmar M, Doody J, Timokhina I, Massague J (1999) A mechanism of repression of TGFbeta/Smad signaling by oncogenic Ras. Genes Dev 13:804–816PubMedCrossRefGoogle Scholar
  26. 26.
    Engel ME, McDonnell MA, Law BK, Moses HL (1999) Interdependent SMAD and JNK signaling in transforming growth factor-beta-mediated transcription. J Biol Chem 274:37413–37420PubMedCrossRefGoogle Scholar
  27. 27.
    Matsuura I, Denissova NG, Wang G, He D, Long J, Liu F (2004) Cyclin-dependent kinases regulate the antiproliferative function of Smads. Nature 430:226–231PubMedCrossRefGoogle Scholar
  28. 28.
    Yakymovych I, Ten Dijke P, Heldin CH, Souchelnytskyi S (2001) Regulation of Smad signaling by protein kinase C. FASEB J 15:553–555PubMedGoogle Scholar
  29. 29.
    Wicks SJ, Lui S, Abdel-Wahab N, Mason RM, Chantry A (2000) Inactivation of smad-transforming growth factor beta signaling by Ca(2+)-calmodulin-dependent protein kinase II. Mol Cell Biol 20:8103–8111PubMedCrossRefGoogle Scholar
  30. 30.
    Ben Aziz-Aloya R, Seidman S, Timberg R, Sternfeld M, Zakut H, Soreq H (1993) Expression of a human acetylcholinesterase promoter-reporter construct in developing neuromuscular junctions of Xenopus embryos. Proc Natl Acad Sci USA 90:2471–2475PubMedCrossRefGoogle Scholar
  31. 31.
    Su W, Wu J, Ye WY, Zhang XJ (2008) A monoclonal antibody against synaptic AChE: a useful tool for detecting apoptotic cells. Chem Biol Interact 175:101–107PubMedCrossRefGoogle Scholar
  32. 32.
    Guillemin I, Becker M, Ociepka K, Friauf E, Nothwang HG (2005) A subcellular prefractionation protocol for minute amounts of mammalian cell cultures and tissue. Proteomics 5:35–45PubMedCrossRefGoogle Scholar
  33. 33.
    Chow EK, O’Connell RM, Schilling S, Wang XF, Fu XY, Cheng G (2005) TLR agonists regulate PDGF-B production and cell proliferation through TGF-beta/type I IFN crosstalk. EMBO J 24:4071–4081PubMedCrossRefGoogle Scholar
  34. 34.
    Tzachanis D, Li L, Lafuente EM, Berezovskaya A, Freeman GJ, Boussiotis VA (2007) Twisted gastrulation (Tsg) is regulated by Tob and enhances TGF-beta signaling in activated T lymphocytes. Blood 109:2944–2952PubMedGoogle Scholar
  35. 35.
    Zhang Y, Feng X, We R, Derynck R (1996) Receptor-associated Mad homologues synergize as effectors of the TGF-beta response. Nature 383:168–172PubMedCrossRefGoogle Scholar
  36. 36.
    Toiber D, Soreq H (2005) Cellular stress reactions as putative cholinergic links in Alzheimer’s disease. Neurochem Res 30:909–919PubMedCrossRefGoogle Scholar
  37. 37.
    Meshorer E, Toiber D, Zurel D, Sahly I, Dori A, Cagnano E, Schreiber L, Grisaru D, Tronche F, Soreq H (2004) Combinatorial complexity of 5′ alternative acetylcholinesterase transcripts and protein products. J Biol Chem 279:29740–29751PubMedCrossRefGoogle Scholar
  38. 38.
    Meshorer E, Soreq H (2006) Virtues and woes of AChE alternative splicing in stress-related neuropathologies. Trends Neurosci 29:216–224PubMedCrossRefGoogle Scholar
  39. 39.
    Landfield PW (1987) ‘Increased calcium-current’ hypothesis of brain aging. Neurobiol Aging 8:346–347PubMedCrossRefGoogle Scholar
  40. 40.
    Disterhoft JF, Moyer JR Jr, Thompson LT (1994) The calcium rationale in aging and Alzheimer’s disease. Evidence from an animal model of normal aging. Ann N Y Acad Sci 747:382–406PubMedCrossRefGoogle Scholar
  41. 41.
    Gibson GE, Zhang H, Toral-Barza L, Szolosi S, Tofel-Grehl B (1996) Calcium stores in cultured fibroblasts and their changes with Alzheimer’s disease. Biochim Biophys Acta 1316:71–77PubMedGoogle Scholar
  42. 42.
    LaFerla FM (2002) Calcium dyshomeostasis and intracellular signalling in Alzheimer’s disease. Nat Rev Neurosci 3:862–872PubMedCrossRefGoogle Scholar
  43. 43.
    Waser M, Mesaeli N, Spencer C, Michalak M (1997) Regulation of calreticulin gene expression by calcium. J Cell Biol 138:547–557PubMedCrossRefGoogle Scholar
  44. 44.
    Bartlett JD, Luethy JD, Carlson SG, Sollott SJ, Holbrook NJ (1992) Calcium ionophore A23187 induces expression of the growth arrest and DNA damage inducible CCAAT/enhancer-binding protein (C/EBP)-related gene, gadd153. Ca2+ increases transcriptional activity and mRNA stability. J Biol Chem 267:20465–20470PubMedGoogle Scholar
  45. 45.
    Jackisch C, Hahm HA, Tombal B, McCloskey D, Butash K, Davidson NE, Denmeade SR (2000) Delayed micromolar elevation in intracellular calcium precedes induction of apoptosis in thapsigargin-treated breast cancer cells. Clin Cancer Res 6:2844–2850PubMedGoogle Scholar
  46. 46.
    McColl KS, He H, Zhong H, Whitacre CM, Berger NA, Distelhorst CW (1998) Apoptosis induction by the glucocorticoid hormone dexamethasone and the calcium-ATPase inhibitor thapsigargin involves Bc1–2 regulated caspase activation. Mol Cell Endocrinol 139:229–238PubMedCrossRefGoogle Scholar
  47. 47.
    Kaneko Y, Tsukamoto A (1994) Thapsigargin-induced persistent intracellular calcium pool depletion and apoptosis in human hepatoma cells. Cancer Lett 79:147–155PubMedCrossRefGoogle Scholar
  48. 48.
    Chen F, Ogawa K, Liu X, Stringfield TM, Chen Y (2002) Repression of Smad2 and Smad3 transactivating activity by association with a novel splice variant of CCAAT-binding factor C subunit. Biochem J 364:571–577PubMedCrossRefGoogle Scholar
  49. 49.
    Davies P, Maloney AJ (1976) Selective loss of central cholinergic neurons in Alzheimer’s disease. Lancet 2:1403PubMedCrossRefGoogle Scholar
  50. 50.
    Cerpa W, Dinamarca MC, Inestrosa NC (2008) Structure-function implications in Alzheimer’s disease: effect of Abeta oligomers at central synapses. Curr Alzheimer Res 5:233–243PubMedCrossRefGoogle Scholar
  51. 51.
    Pakaski M, Kalman J (2008) Interactions between the amyloid and cholinergic mechanisms in Alzheimer’s disease. Neurochem Int 53:103–111PubMedCrossRefGoogle Scholar
  52. 52.
    Inestrosa NC, Urra S, Colombres M (2004) Acetylcholinesterase (AChE)–amyloid-beta-peptide complexes in Alzheimer’s disease. The Wnt signaling pathway. Curr Alzheimer Res 1:249–254PubMedCrossRefGoogle Scholar
  53. 53.
    Jean L, Thomas B, Tahiri-Alaoui A, Shaw M, Vaux DJ (2007) Heterologous amyloid seeding: revisiting the role of acetylcholinesterase in Alzheimer’s disease. PLoS ONE 2:e652PubMedCrossRefGoogle Scholar
  54. 54.
    Dinamarca MC, Arrazola M, Toledo E, Cerpa WF, Hancke J, Inestrosa NC (2008) Release of acetylcholinesterase (AChE) from beta-amyloid plaques assemblies improves the spatial memory impairments in APP-transgenic mice. Chem Biol Interact 175:142–149PubMedCrossRefGoogle Scholar
  55. 55.
    Recanatini M, Valenti P (2004) Acetylcholinesterase inhibitors as a starting point towards improved Alzheimer’s disease therapeutics. Curr Pharm Des 10:3157–3166PubMedCrossRefGoogle Scholar
  56. 56.
    Loizzo MR, Tundis R, Menichini F, Menichini F (2008) Natural products and their derivatives as cholinesterase inhibitors in the treatment of neurodegenerative disorders: an update. Curr Med Chem 15:1209–1228PubMedCrossRefGoogle Scholar
  57. 57.
    Ueberham U, Ueberham E, Gruschka H, Arendt T (2006) Altered subcellular location of phosphorylated Smads in Alzheimer’s disease. Eur J NeuroSci 24:2327–2334PubMedCrossRefGoogle Scholar
  58. 58.
    Burton T, Liang B, Dibrov A, Amara F (2002) Transforming growth factor-beta-induced transcription of the Alzheimer beta-amyloid precursor protein gene involves interaction between the CTCF-complex and Smads. Biochem Biophys Res Commun 295:713–723PubMedCrossRefGoogle Scholar
  59. 59.
    Docagne F, Gabriel C, Lebeurrier N, Lesne S, Hommet Y, Plawinski L, Mackenzie ET, Vivien D (2004) Sp1 and Smad transcription factors co-operate to mediate TGF-beta-dependent activation of amyloid-beta precursor protein gene transcription. Biochem J 383:393–399PubMedCrossRefGoogle Scholar
  60. 60.
    Ulrich J, Meier-Ruge W, Probst A, Meier E, Ipsen S (1990) Senile plaques: staining for acetylcholinesterase and A4 protein: a comparative study in the hippocampus and entorhinal cortex. Acta Neuropathol 80:624–628PubMedCrossRefGoogle Scholar

Copyright information

© Birkhäuser Verlag, Basel/Switzerland 2009

Authors and Affiliations

  • Wei Gao
    • 1
  • Hui Zhu
    • 1
  • Jing-Ya Zhang
    • 1
  • Xue-Jun Zhang
    • 1
  1. 1.Laboratory of Molecular Cell Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological SciencesChinese Academy of SciencesShanghaiChina

Personalised recommendations