Molecular and Cellular Biochemistry

, Volume 199, Issue 1–2, pp 139–147 | Cite as

Specific down-regulation of annexin II expression in human cells interferes with cell proliferation

  • Yangping Chiang
  • Angie Rizzino
  • Zita A. Sibenaller
  • Marc S. Wold
  • Jamboor K. Vishwanatha


The protein-tyrosine kinase substrate annexin II is a growth regulated gene whose expression is increased in several human cancers. While the precise function of this protein is not understood, annexin II is proposed to be involved in multiple physiological activities, including DNA synthesis and cell proliferation. Targeted disruption of the annexin II gene affects calcium signaling, tyrosine phosphorylation and apoptosis, indicating the important physiological role of this protein. We used a transient co-transfection assay to regulate annexin II expression in human HeLa, 293 and 293T cells, and measured the effects of annexin II down regulation on DNA synthesis and proliferation. Transfection of cells with an antisense annexin II vector results in inhibition of cell division and proliferation, with concomitant reduction in annexin II message and protein levels. Cellular DNA synthesis is significantly reduced in antisense transfected cells. Replication extracts made from antisense transfected cells have significantly reduced efficiency to support SV40 in vitro DNA replication, while the extracts made from sense transfected cells are fully capable of replication. Our results indicate an important role of annexin II in cellular DNA synthesis and cell proliferation.

annexin II DNA synthesis antisense Simian Virus 40 cell proliferation 


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  1. 1.
    Zokas L, Glenney J Jr: The calpactin light chain is tightly linked to the cytoskeletal form of calpactin I: Studies using monoclonal antibodies to calpactin subunits. J Cell Biol 105: 2111–2121, 1987Google Scholar
  2. 2.
    Kumble KD, Vishwanatha JK: Immunoelectron microscopic analysis of the intracellular distribution of primer recognition proteins, annexin 2 and phosphoglycerate kinase, in normal and transformed cells. J Cell Sci 99: 751–758, 1991Google Scholar
  3. 3.
    Vishwanatha JK, Jindal HK, Davis RG: The role of primer recognition proteins in DNA replication: Association with nuclear matrix in HeLa cells. J Cell Sci 101: 25–34, 1992Google Scholar
  4. 4.
    Tressler RJ, Updyke TV, Yeatman T, Nicolson GL: Extracellular annexin II is associated with divalent cation-dependent tumor cellendothelial cell adhesion of metastatic RAW117 large-cell lymphoma cells. J Cell Biochem 53: 265–276, 1993Google Scholar
  5. 5.
    Keutzer JC, Hirschhorn RR: The growth-regulated gene 1B6 is identified as the heavy chain of calpactin I. Exp Cell Res 188: 153–159, 1990Google Scholar
  6. 6.
    Vellucci VF, Germino FJ, Reiss M: Cloning of putative growth regulatory genes from primary human keratinocytes by subtractive hybridization. Gene 166: 213–220, 1995Google Scholar
  7. 7.
    Chiang Y, Schneiderman MH, Vishwanatha JK: Annexin II expression is regulated during mammalian cell cycle. Cancer Res 53: 6017–6021, 1993Google Scholar
  8. 8.
    Cole SPC, Pinkoski MJ, Bhardwaj G, Deeley RG: Elevated expression of Annexin II (Lipocortin II, p36) in a multidrug resistant small cell lung cancer cell line. Br J Cancer 65: 498–502, 1992Google Scholar
  9. 9.
    Frohlich M, Motte P, Galvin K, Takahashi H, Wands J, Ozturk M: Enhanced expression of the protein kinase substrate p36 in human hepatocellular carcinoma. Mol Cell Biol 10: 3216–3223, 1990Google Scholar
  10. 10.
    Kumble KD, Hirota M, Pour PM, Vishwanatha JK: Enhanced levels of annexins in pancreatic carcinoma cells of Syrian hamsters and their intrapancreatic allografts. Cancer Res 52: 163–167, 1992Google Scholar
  11. 11.
    Reeves SA, Chavez-Kappel C, Davis R, Rosenblum M, Israel MA: Developmental regulation of annexin II (lipocortin 2) in human brain and expression in high grade glioma. Cancer Res 52: 6871–6876, 1992Google Scholar
  12. 12.
    Vishwanatha JK, Chiang Y, Kumble KD, Hollingsworth MA, Pour PM: Enhanced expression of annexin II in human pancreatic carcinoma cells and primary pancreatic cancers. Carcinogenesis 14: 2575–2579, 1993Google Scholar
  13. 13.
    Jindal HK, Chaney WG, Anderson CW, Davis RG, Vishwanatha JK: The protein-tyrosine kinase substrate, calpactin I heavy chain (p36), is part of the primer recognition protein complex that interacts with DNA polymerase α. J Biol Chem 266: 5169–5176, 1991Google Scholar
  14. 14.
    Jindal HK, Vishwanatha JK: Purification and characterization of primer recognition proteins from HeLa cells. Biochemistry 29: 4767–4773, 1990Google Scholar
  15. 15.
    Vishwanatha JK, Kumble S: Involvement of annexin II in DNA replication: Evidence from cell-free extracts of Xenopus eggs. J Cell Sci 105: 533–540, 1993Google Scholar
  16. 16.
    Ma YG, Rosfjord E, Huebert C, Wilder P, Tiesman J, Kelly D, Rizzino A: Transcriptional regulation of the murine k-FGF gene in embryonic cell lines. Dev Biol 154: 45–54, 1992Google Scholar
  17. 17.
    Rosner MH, Vigano MA, Ozato K, Timmons PM, Poirier F, Rigby PW, Staudt LM: A POU-domain transcription factor in early stem cells and germ cells of the mammalian embryo. Nature 345: 686–692, 1990Google Scholar
  18. 18.
    Minucci S, Botquin V, Yeom YI, Dey A, Sylvester I, Z and DJ, Ohbo K, Ozato K, Scholer HR: Retinoic acid-mediated down-regulation of Oct3/4 coincides with the loss of promoter occupancy in vivo. Embo J 15: 888–899, 1996Google Scholar
  19. 19.
    Paabo S, Weber F, Nilsson T, Schaffner W, Peterson PA: Structural and functional dissection of an MHC class I antigen-binding adenovirus glycoprotein. Embo J 5: 1921–1927, 1986Google Scholar
  20. 20.
    Wold MS, Li JJ, Kelly TJ: Initiation of simian virus 40 DNA replication in vitro: Large-tumor-antigen-and origin-dependent unwinding of the template. Proc Natl Acad Sci USA 84: 3643–3647, 1987Google Scholar
  21. 21.
    Henricksen LA, Umbricht CB, Wold MS: Recombinant replication protein A: Expression, complex formation and functional characterization. J Biol Chem 269: 11121–11132, 1994Google Scholar
  22. 22.
    Hendricks TL, Darlington GJ: Inhibition of cell proliferation by C/EBP alpha occurs in many cell types, does not require the presence of p53 or Rb, and is not affected by large T-antigen. Nucl Acids Res 23: 4726–4733, 1995Google Scholar
  23. 23.
    McGowan CH, Russell P: Human wee1 kinase inhibits cell division by phosphorylating p34cdc2 exclusively on Tyr 15. Embo J 12: 75–85, 1993Google Scholar
  24. 24.
    Wakita K, Lord JM, Tokuhisa T: A method for detecting the expression of a toxic gene in cultured cells. Anal Biochem 198: 224–227, 1991Google Scholar
  25. 25.
    Friedman RL, Manly SP, McMahon M, Kerr IM, Stark GR: Transcriptional and posttranscriptional regulation of interferon-induced gene expression in human cells. Cell 38: 745–755, 1984Google Scholar
  26. 26.
    Numa F, Hirabayashi K, Tsunaga N, Kato H, O'Rourke K, Shao H, Stechmann LC, Varani J, Rapraeger A, Dixit VM: Elevated levels of syndecan-1 expression confer potent serum-dependent growth in human 293T cells. Cancer Res 55: 4676–4680, 1995Google Scholar
  27. 27.
    Koda T, Hasan S, Sasaki A, Arimura Y, Kakinuma M: Regulatory sequences required for hst-1 expression in embryonal carcinoma cells. FEBS Lett 342: 71–75, 1994Google Scholar
  28. 28.
    Pear WS, Nolan GP, Scott ML, Baltimore D: Production of high-titer helper-free retroviruses by transient transfection. Proc Natl Acad Sci USA 90: 8392–8396, 1993Google Scholar
  29. 29.
    Klee CB: Ca2+-dependent phospholipid-(and membrane-) binding proteins. Biochemistry 27: 6645–6653, 1988Google Scholar
  30. 30.
    Vishwanatha JK, Coughlin SA, Wesolowski-Owen M, Baril EF: A multiprotein form of DNA polymerase α from HeLa cells: Resolution of its associated catalytic activities. J Biol Chem 261: 6619–6628, 1986Google Scholar

Copyright information

© Kluwer Academic Publishers 1999

Authors and Affiliations

  • Yangping Chiang
    • 1
  • Angie Rizzino
    • 2
  • Zita A. Sibenaller
    • 3
  • Marc S. Wold
    • 3
  • Jamboor K. Vishwanatha
    • 1
    • 2
  1. 1.Department of Biochemistry and Molecular BiologyUniversity of Nebraska Medical CenterOmahaUSA
  2. 2.Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical CenterOmahaUSA
  3. 3.Department of BiochemistryUniversity of Iowa College of MedicieIowa CityUSA

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