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Tyrosine phosphorylation of a ∼30 kd protein precedes αvβ3 integrin-signaled endothelial cell spreading and motility on matrix proteins

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Pathology & Oncology Research

Abstract

A microvascular endothelial cell line (CD clone 4) isolated from murine lung adheres to and spreads well on fibronectin, vitronectin, and fibrinogen, but poorly on collagen type IV and laminin. Ligating cell surface αv, β3, α4, α5, or β1 integrin receptors with monospecific antibodies promoted a dramatic cell spreading and motility on vitronectin or collagen IV. Antibodies directed to other adhesion molecules, including αIIb, PECAM-1, and P-selectin were ineffective. Ligation with monoclonal anti-αv or -β3, but not -α4, -α5, or -β1 antibodies, induced a rapid, and dose-dependent tyrosine phosphorylation of a ∼30 kD protein, which preceded CD clone 4 endothelial cell spreading and motility and was partially inhibited by genistein and completely inhibited by BAPTA. All other antibodies tested did not induce the tyrosine phosphorylation of the 30 kD protein as well as cell spreading and motility. The present results suggest that β1 and β3 integrins employ different biochemical mechanisms in signaling endothelial cell spreading and motility and that the tyrosine phosphorylation of the 30 kD protein (and probably other proteins) may play an important role in signaling β3 integrin-mediated endothelial cell interaction with other cells (e.g., tumor cells) and extracellular matrix.

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Abbreviations

Ab:

antibody

BAPTA-AM:

bis(2-aminophenoxy)-ethane-N, N, N N’-tetraacetate, acetoxymethyl ester

DMEM:

Dulbecco’s Minimum Essential Medium

FAK:

focal adhesion kinase

mAb:

monoclonal antibody

pAb:

polyclonal antibody

PKC:

protein kinase C

SKM:

subendothelial matrix

Vn:

vitronectin

References

  1. Albelda SM: Endothelial and epithelial cell adhesion molecules. Am J Respir Cell Mol Biol 8: 195–203. 1991.

    Google Scholar 

  2. Honn KV and Tang DG: Adhesion molecules and cancer cell interaction with endothelium and subendothelial matrix. Cancer Metastasis Rev 11: 353–375. 1992.

    Article  PubMed  CAS  Google Scholar 

  3. Massia SP and Hubbel JA: Vascular endothelial cell adhesion and spreading promoted by the peptide REDV of IIICS region of plasma fibronectin is mediated by integrin alpha 4 beta I. J Biol Chem 267: 1 4019–14026. 1992.

    Google Scholar 

  4. Tang DG, Chen Y, Diglio CA and Honn KV: Protein kinase Cdependent effects of 12(S)-HETE on endothelial cell vitronectin and fibronectin receptor. J Cell Biol 121: 689–704. 1993.

    Article  PubMed  CAS  Google Scholar 

  5. Lampugnani MG, Resnati M, Dejana E and Marchisio PC: The role of integrins in the maintenance of endothelial cell monolayer integrity. J Cell Biol 112: 479–490. 1991.

    Article  PubMed  CAS  Google Scholar 

  6. Lafrenie RM, Podor TJ, Buchanan MR and Orr FW: Upregulated biosynthesis and expression of endothelial cell vitronectin receptor enhances cancer cell adhesion. Cancer Res 52: 2202–2208. 1992.

    PubMed  CAS  Google Scholar 

  7. Tang DG, Grossi IM, Chen Y, Diglio CA and Honn KV: 12(S)-HETE promotes tumor cell adhesion by increasing surface expression of αvβ3 integrins on endothelial cells. Int J Cancer 54: 102–111. 1993.

    Article  PubMed  CAS  Google Scholar 

  8. Tang DG, Diglio CA and Honn KV: Activation of microvascular endothelialium by 12(S)-HETE leads to enhanced tumor cell adhesion via upregulation of surface expression of αvβ3 integrin: A post-transcriptional, PKC-and cytoskeleton-dependent process. Cancer Res 54: 1119–1129. 1994.

    PubMed  CAS  Google Scholar 

  9. Hynes RO: Integrins: Versatility, modulation. and signaling in cell adhesion. Cell 69: 11–25. 1992.

    Article  PubMed  CAS  Google Scholar 

  10. Schwartz MA, Ingber D, Lawrence M, Springer TM and Lechene C: Multiple integrins share the ability to induce elevation of intracellular pH. Exp Cell Res 195: 533–535. 1991.

    Article  PubMed  CAS  Google Scholar 

  11. Romer LH, McLean N, Turner CE and Burridge K: Tyrosine kinase activity, cytoskeletal organization, and motility in human vascular endothelial cells. Mol. Biol. Cell 5: 349–361. 1994.

    PubMed  CAS  Google Scholar 

  12. Schwartz MA: Spreading of human endothelial cells on fibronectin or vitronectin triggers elevation of intracellular free calcium. J Cell Biol 120: 1003–1010, 1993.

    Article  PubMed  CAS  Google Scholar 

  13. Leavesley DI, Schwartz MA, Rosenfeld M and Cheresh DA: Integrin β1 and β3-mediated endothelial cell migration is triggered through distinct signaling mechanisms. J Cell Biol 121: 163–170. 1993.

    Article  PubMed  CAS  Google Scholar 

  14. Bates RC, Buret A, van Helden DF, Horton MA and Burns GF: Apoptosis induced inhibition of intercellular contact. J Cell Biol 125: 403–415. 1994.

    Article  PubMed  CAS  Google Scholar 

  15. Brooks P, Clark RAF and Cheresh DA: Requirement of vascular integrin αvβ3 for angiogenesis. Science 264: 569–571. 1994.

    Article  PubMed  CAS  Google Scholar 

  16. Juliano RL and Haskill S: Signal transduction from extracellular matrix J. Cell Biol. 120: 577–585. 1993.

    Article  PubMed  CAS  Google Scholar 

  17. Re F, Zanetti A, Sironi M, Polentarutti N, Lanfrancone L, Dejana E and Colotta F: Inhibition of anchorage-dependent cell spreading triggers apoptosis in cultured human endothelial cells. J Cell Biol 127: 537–546. 1994.

    Article  PubMed  CAS  Google Scholar 

  18. Guan JL and Shalloway D: Regulation of focal adhesion associated protein tyrosine kinase by both cellular adhesion and oncogenic transformation. Nature 358: 690–692, 1992.

    Article  PubMed  CAS  Google Scholar 

  19. Kornberg LJ, Earp HS, Turner CE, Prockop C and Juliano RL: Signal transduction by integrins: increased protein tyrosine phosphorylation caused by clustering of beta integrins. Proc Natl Acad Sci USA 88: 8392–8396. 1991.

    Article  PubMed  CAS  Google Scholar 

  20. Burridge K, Turner CE and Romer LH: Tyrosine phosphorylation of paxillin and ppl25EAR accompanies cell adhesion to extracellular matrix: a role in cytoskeletal assembly. J Cell Biol 119: 893–903. 1992.

    Article  PubMed  CAS  Google Scholar 

  21. Bockholt SM and Burridge K: Cell spreading on extracellular matrix proteins induces tyrosine phosphorylation of tensin. J Biol Chem 268: 14565–14567. 1993.

    PubMed  CAS  Google Scholar 

  22. Tang DG, Diglio CA, Bazaz R and Honn KV: Transcriptional activation of endothelial cell integrin αv by protein kinase C activator 12(S)-HETE. J Cell Sci 108: 2629–2644. 1995.

    PubMed  Google Scholar 

  23. Timar J, Silletti S, Bazaz R, Raz A and Honn KV: Regulation of melanoma cell motility by the lipoxygenase metabolite 12(S)-HETE. Int J Cancer 55: 1003–1010. 1993.

    Article  PubMed  CAS  Google Scholar 

  24. Freedman AS, Rhynhat K, Nojima Y, Svahn J, Eliseo L, Benjamin CD, Morimoto C and Vivier E: Stimulation of protein tyrosine phopshorylation in human B cells after ligation of the β1 integrin VLA4.J Immunol 150: 1645–1652. 1993.

    PubMed  CAS  Google Scholar 

  25. Lin TH, Yurochko A, Kornherg L, Morris J, Walker JJ, Haskill S and Juliano RL: The role of protein tyrosine phosphorylation in integrin-mediated gene induction in monocytes. J Cell Biol 126: 1585–1593. 1994.

    Article  PubMed  CAS  Google Scholar 

  26. Siaastad MD, Angres B, Lewis RS and Nelson WJ: Feedback regulation of cell-substratum adhesion by integrin-mediated intracellular Ca2+ signaling. Proc Natl Acad Sci USA 91: 8214–8218.1994.

    Article  Google Scholar 

  27. Clyman RL, Maury L, and Kramer RH: β1 and β3 integrins have different roles in the adhesion and migration of vascular smooth muscle cells on extracellular matrix. Exp Cell Res 200: 272–284. 1992.

    Article  PubMed  CAS  Google Scholar 

  28. Shankar G, Davison I, Helfrich MH, Mason WT, and Horton MA: Integrin-receptor-mediated mobilization of intracellular calcium in rat osteoclasts. J Cell Sci 105: 61–68. 1993.

    PubMed  CAS  Google Scholar 

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Authors and Affiliations

  1. Department of Radiation Oncology, Wayne State University, 431 Chemistry, 48202, Detroit, Ml, USA

    Dean G. Tang & Kenneth V Honn

  2. Department of Radiation Pathology, Gershenson Radiation Oncology Center, Detroit

    Clement A Diglio

  3. Department of Radiation Chemistry, Harper Hospital, Detroit, USA

    Kenneth V Honn

Authors
  1. Dean G. Tang
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  2. Clement A Diglio
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  3. Kenneth V Honn
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Tang, D.G., Diglio, C.A. & Honn, K.V. Tyrosine phosphorylation of a ∼30 kd protein precedes αvβ3 integrin-signaled endothelial cell spreading and motility on matrix proteins. Pathol. Oncol. Res. 2, 21–29 (1996). https://doi.org/10.1007/BF02893943

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  • DOI: https://doi.org/10.1007/BF02893943

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