Use of an immobilized monoclonal antibody to examine integrin α5β1 signaling independent of cell spreading

被引：6
作者：
Bao W. [1 ]
Strömblad S. [1 ]
机构：
[1] Karolinska Institutet, Dept. of Microbiol. Pathol./Immunol., Huddinge University Hospital, Huddinge
来源：
Biological Procedures Online | 2002年 / 4卷 / 1期
关键词：
Antibodies; Cell adhesion; Fibronectins; Integrins; Monoclonal; Protein kinases;
D O I：
10.1251/bpo37
中图分类号：
学科分类号：
摘要：
Cell attachment to the extracellular matrix (ECM) engages integrin signaling into the cell, but part of the signaling response also stem from cell spreading (3). To analyze specific integrin signaling-mediated responses independent of cell spreading, we developed a method engaging integrin signaling by use of an immobilized anti-integrin monoclonal antibody (mab) directed against the fibronectin (FN) receptor integrin α5β1. ECV 304 cells were plated onto FN or immobilized mab JBS5 (anti-integrin α5β1) or onto poly-L-lysin (P-L-L), which mediates integrin-independent attachment. Cells attached and spread on FN, while cells on JBS5 or P-L-L attached but did not spread. Importantly, plating onto FN or mab JBS5 gave rise to identical integrin-induced responses, including a down-regulation of the cyclin-dependent kinase (Cdk2) inhibitors p21CIP1 and p27KIP1, while attachment to P-L-L did not. We conclude that engagement of the FN-receptor integrin α5β1 induces integrin signaling regulating the Cdk2-inhibitors independent of cell spreading and present a method for how integrin signaling can be analyzed separate from the effects of cell spreading.
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收藏
页码：81 / 87
页数：6
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共 12 条
  • [1] Bao W.J., Thullberg M., Zhang H.Q., Onischenko A., Stormblad S., Cell attachment to the extracellular matrix induces proteasomal degradation of p21<sup>CIP1</sup> via Cdc42/Rac1 signaling, Mol. Cell Biol., 22, pp. 4587-4597, (2002)
  • [2] Brown J., Reading S.J., Jones S., Fitchett C.J., Howl J., Martin A., Longland C., Michelangeli F., Dubrova Y.E., Brown C.A., Critical evaluation of ECV 304 as a human endothelial cell model defined by genetic analysis and functional responses: A comparison with the human bladder cancer derived epithelial cell line T24/83, Lab. Invest., 80, pp. 37-45, (2000)
  • [3] Chen C.S., Marksich M., Huang S., Whitesides G.M., Ingber D.E., Geometric control of cell life and death, Science, 276, pp. 1425-1428, (1997)
  • [4] Chen Q., Lin T.H., Der C.J., Juliano R.L., Integrin-mediated activation of mitogen-activated protein (MAP) or extracellular signal-related kinase kinase (MEK) and kinase is independent of Ras, J. Biol. Chem., 271, pp. 18122-18127, (1996)
  • [5] Giancotti F.G., Ruoslahti E., Integrin signaling, Science, 285, pp. 1028-1032, (1999)
  • [6] Holmes E., Engvall E., Determination of integrins on cells by cell adhesion to antibodies, Anal. Biochem., 214, pp. 100-105, (1993)
  • [7] Hynes R.O., Integrins: Versatility, modulation, and signaling in cell adhesion, Cell, 69, pp. 11-25, (1992)
  • [8] Leavesley D.I., Schwartz M.A., Rosenfeld M., Cheresh D.A., Integrin beta 1- and beta 3 mediated endothelial cell migration is triggered through distinct signaling mechanisms, J. Cell Biol., 121, pp. 163-170, (1993)
  • [9] Plow E.F., Hass T.A., Zhang L., Loftus J., Smith J.W., Ligand binding to integrins, J. Biol. Chem., 275, pp. 21785-21788, (2000)
  • [10] Ruoslahti E., Fibronectin and its receptors, Annu. Rev. Biochem., 57, pp. 375-413, (1988)
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