Skip to main content
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1987 Oct 1;105(4):1867–1872. doi: 10.1083/jcb.105.4.1867

Chemotaxis of 3T3 and SV3T3 cells to fibronectin is mediated through the cell-attachment site in fibronectin and a fibronectin cell surface receptor

PMCID: PMC2114639  PMID: 2959668

Abstract

Fibronectin (FN) is a multidomain extracellular matrix protein that induces attachment and chemotactic migration of fibroblastic cells. In this study we analyzed the molecular determinants involved in the FN- induced chemotactic migration of normal and SV40-transformed 3T3 cells. Two different monoclonal antibodies to the cell-binding site of FN blocked chemotaxis to a 140-kD FN fragment (Ca 140) containing the cell- binding domain. A monoclonal antibody to a determinant distant from the cell-binding site did not affect chemotaxis. A synthetic tetrapeptide, RGDS, which represents the major cell-attachment sequence, was able to compete with FN and the Ca 140 fragment in chemotaxis assays, but this peptide itself had no significant chemotactic activity. A larger peptide encompassing this sequence, GRGDSP, was chemotactic, while the peptide GRGESP, where a glutamic acid residue was substituted for aspartic acid, was inactive. Chemotactic migration could be prevented in a dose-dependent manner by a rabbit polyclonal antiserum to a 140-kD cell surface FN receptor. This antibody was more effective on normal than on transformed 3T3 cells. Neither the anti-FN receptor antiserum nor a monoclonal antibody to the cell-binding site of FN blocked migration induced by another potent chemoattractant, platelet-derived growth factor. These data indicate that FN-induced chemotaxis of 3T3 and SV3T3 cells is mediated via the RGDS cell-attachment site of FN and the 140-kD cell surface FN receptor. The interaction is specific and can be altered by transformation.

Full Text

The Full Text of this article is available as a PDF (721.0 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Akiyama S. K., Yamada S. S., Yamada K. M. Characterization of a 140-kD avian cell surface antigen as a fibronectin-binding molecule. J Cell Biol. 1986 Feb;102(2):442–448. doi: 10.1083/jcb.102.2.442. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Albini A., Adelmann-Grill B. C., Müller P. K. Fibroblast chemotaxis. Coll Relat Res. 1985 Jun;5(3):283–296. doi: 10.1016/s0174-173x(85)80018-2. [DOI] [PubMed] [Google Scholar]
  3. Albini A., Allavena G., Richter H., Pontz B. F., Parodi S., Santi L. SV40 transformed fibroblasts recognize the same 140 kD fibronectin chemotactic fragment as non-transformed cells. Experientia. 1985 Dec 15;41(12):1591–1593. doi: 10.1007/BF01964822. [DOI] [PubMed] [Google Scholar]
  4. Albini A., Richter H., Pontz B. F. Localization of the chemotactic domain in fibronectin. FEBS Lett. 1983 Jun 13;156(2):222–226. doi: 10.1016/0014-5793(83)80500-6. [DOI] [PubMed] [Google Scholar]
  5. Aswanikumar S., Corcoran B., Schiffmann E., Day A. R., Freer R. J., Showell H. J., Becker E. L. Demonstration of a receptor on rabbit neutrophils for chemotactic peptides. Biochem Biophys Res Commun. 1977 Jan 24;74(2):810–817. doi: 10.1016/0006-291x(77)90375-8. [DOI] [PubMed] [Google Scholar]
  6. Betsholtz C., Johnsson A., Heldin C. H., Westermark B., Lind P., Urdea M. S., Eddy R., Shows T. B., Philpott K., Mellor A. L. cDNA sequence and chromosomal localization of human platelet-derived growth factor A-chain and its expression in tumour cell lines. Nature. 1986 Apr 24;320(6064):695–699. doi: 10.1038/320695a0. [DOI] [PubMed] [Google Scholar]
  7. Brown P. J., Juliano R. L. Selective inhibition of fibronectin-mediated cell adhesion by monoclonal antibodies to a cell-surface glycoprotein. Science. 1985 Jun 21;228(4706):1448–1451. doi: 10.1126/science.4012302. [DOI] [PubMed] [Google Scholar]
  8. Chen W. T., Greve J. M., Gottlieb D. I., Singer S. J. Immunocytochemical localization of 140 kD cell adhesion molecules in cultured chicken fibroblasts, and in chicken smooth muscle and intestinal epithelial tissues. J Histochem Cytochem. 1985 Jun;33(6):576–586. doi: 10.1177/33.6.3889142. [DOI] [PubMed] [Google Scholar]
  9. Chiang T. M., Postlethwaite A. E., Beachey E. H., Seyer J. M., Kang A. H. Binding of chemotactic collagen-derived peptides to fibroblasts. The relationship to fibroblast chemotaxis. J Clin Invest. 1978 Nov;62(5):916–922. doi: 10.1172/JCI109219. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Damsky C. H., Knudsen K. A., Bradley D., Buck C. A., Horwitz A. F. Distribution of the cell substratum attachment (CSAT) antigen on myogenic and fibroblastic cells in culture. J Cell Biol. 1985 May;100(5):1528–1539. doi: 10.1083/jcb.100.5.1528. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Duband J. L., Rocher S., Chen W. T., Yamada K. M., Thiery J. P. Cell adhesion and migration in the early vertebrate embryo: location and possible role of the putative fibronectin receptor complex. J Cell Biol. 1986 Jan;102(1):160–178. doi: 10.1083/jcb.102.1.160. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Dziadek M., Richter H., Schachner M., Timpl R. Monoclonal antibodies used as probes for the structural organization of the central region of fibronectin. FEBS Lett. 1983 May 8;155(2):321–325. doi: 10.1016/0014-5793(82)80629-7. [DOI] [PubMed] [Google Scholar]
  13. Gauss-Müller V., Kleinman H. K., Martin G. R., Schiffmann E. Role of attachment factors and attractants in fibroblast chemotaxis. J Lab Clin Med. 1980 Dec;96(6):1071–1080. [PubMed] [Google Scholar]
  14. Giancotti F. G., Comoglio P. M., Tarone G. A 135,000 molecular weight plasma membrane glycoprotein involved in fibronectin-mediated cell adhesion. Immunofluorescence localization in normal and RSV-transformed fibroblasts. Exp Cell Res. 1986 Mar;163(1):47–62. doi: 10.1016/0014-4827(86)90557-4. [DOI] [PubMed] [Google Scholar]
  15. Giancotti F. G., Languino L. R., Zanetti A., Peri G., Tarone G., Dejana E. Platelets express a membrane protein complex immunologically related to the fibroblast fibronectin receptor and distinct from GPIIb/IIIa. Blood. 1987 May;69(5):1535–1538. [PubMed] [Google Scholar]
  16. Giancotti F. G., Tarone G., Knudsen K., Damsky C., Comoglio P. M. Cleavage of a 135 kD cell surface glycoprotein correlates with loss of fibroblast adhesion to fibronectin. Exp Cell Res. 1985 Jan;156(1):182–190. doi: 10.1016/0014-4827(85)90272-1. [DOI] [PubMed] [Google Scholar]
  17. Greve J. M., Gottlieb D. I. Monoclonal antibodies which alter the morphology of cultured chick myogenic cells. J Cell Biochem. 1982;18(2):221–229. doi: 10.1002/jcb.1982.240180209. [DOI] [PubMed] [Google Scholar]
  18. Hasegawa T., Hasegawa E., Chen W. T., Yamada K. M. Characterization of a membrane-associated glycoprotein complex implicated in cell adhesion to fibronectin. J Cell Biochem. 1985;28(4):307–318. doi: 10.1002/jcb.240280409. [DOI] [PubMed] [Google Scholar]
  19. Heldin C. H., Westermark B., Wasteson A. Specific receptors for platelet-derived growth factor on cells derived from connective tissue and glia. Proc Natl Acad Sci U S A. 1981 Jun;78(6):3664–3668. doi: 10.1073/pnas.78.6.3664. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Horwitz A., Duggan K., Greggs R., Decker C., Buck C. The cell substrate attachment (CSAT) antigen has properties of a receptor for laminin and fibronectin. J Cell Biol. 1985 Dec;101(6):2134–2144. doi: 10.1083/jcb.101.6.2134. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Humphries M. J., Akiyama S. K., Komoriya A., Olden K., Yamada K. M. Identification of an alternatively spliced site in human plasma fibronectin that mediates cell type-specific adhesion. J Cell Biol. 1986 Dec;103(6 Pt 2):2637–2647. doi: 10.1083/jcb.103.6.2637. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Hynes R. O., Yamada K. M. Fibronectins: multifunctional modular glycoproteins. J Cell Biol. 1982 Nov;95(2 Pt 1):369–377. doi: 10.1083/jcb.95.2.369. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Lacovara J., Cramer E. B., Quigley J. P. Fibronectin enhancement of directed migration of B16 melanoma cells. Cancer Res. 1984 Apr;44(4):1657–1663. [PubMed] [Google Scholar]
  24. McCarthy J. B., Basara M. L., Palm S. L., Sas D. F., Furcht L. T. The role of cell adhesion proteins--laminin and fibronectin--in the movement of malignant and metastatic cells. Cancer Metastasis Rev. 1985;4(2):125–152. doi: 10.1007/BF00050692. [DOI] [PubMed] [Google Scholar]
  25. McCarthy J. B., Furcht L. T. Laminin and fibronectin promote the haptotactic migration of B16 mouse melanoma cells in vitro. J Cell Biol. 1984 Apr;98(4):1474–1480. doi: 10.1083/jcb.98.4.1474. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. McCarthy J. B., Hagen S. T., Furcht L. T. Human fibronectin contains distinct adhesion- and motility-promoting domains for metastatic melanoma cells. J Cell Biol. 1986 Jan;102(1):179–188. doi: 10.1083/jcb.102.1.179. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Mensing H., Albini A., Krieg T., Pontz B. F., Müller P. K. Enhanced chemotaxis of tumor-derived and virus-transformed cells to fibronectin and fibroblast-conditioned medium. Int J Cancer. 1984 Jan 15;33(1):43–48. doi: 10.1002/ijc.2910330109. [DOI] [PubMed] [Google Scholar]
  28. Mensing H., Pontz B. F., Müller P. K., Gauss-Müller V. A study on fibroblast chemotaxis using fibronectin and conditioned medium as chemoattractants. Eur J Cell Biol. 1983 Jan;29(2):268–273. [PubMed] [Google Scholar]
  29. Neff N. T., Lowrey C., Decker C., Tovar A., Damsky C., Buck C., Horwitz A. F. A monoclonal antibody detaches embryonic skeletal muscle from extracellular matrices. J Cell Biol. 1982 Nov;95(2 Pt 1):654–666. doi: 10.1083/jcb.95.2.654. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Pierschbacher M. D., Hayman E. G., Ruoslahti E. Location of the cell-attachment site in fibronectin with monoclonal antibodies and proteolytic fragments of the molecule. Cell. 1981 Oct;26(2 Pt 2):259–267. doi: 10.1016/0092-8674(81)90308-1. [DOI] [PubMed] [Google Scholar]
  31. Pierschbacher M. D., Hayman E. G., Ruoslahti E. The cell attachment determinant in fibronectin. J Cell Biochem. 1985;28(2):115–126. doi: 10.1002/jcb.240280205. [DOI] [PubMed] [Google Scholar]
  32. Pierschbacher M. D., Ruoslahti E. Cell attachment activity of fibronectin can be duplicated by small synthetic fragments of the molecule. Nature. 1984 May 3;309(5963):30–33. doi: 10.1038/309030a0. [DOI] [PubMed] [Google Scholar]
  33. Pierschbacher M. D., Ruoslahti E. Variants of the cell recognition site of fibronectin that retain attachment-promoting activity. Proc Natl Acad Sci U S A. 1984 Oct;81(19):5985–5988. doi: 10.1073/pnas.81.19.5985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Postlethwaite A. E., Keski-Oja J., Balian G., Kang A. H. Induction of fibroblast chemotaxis by fibronectin. Localization of the chemotactic region to a 140,000-molecular weight non-gelatin-binding fragment. J Exp Med. 1981 Feb 1;153(2):494–499. doi: 10.1084/jem.153.2.494. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Pytela R., Pierschbacher M. D., Ruoslahti E. Identification and isolation of a 140 kd cell surface glycoprotein with properties expected of a fibronectin receptor. Cell. 1985 Jan;40(1):191–198. doi: 10.1016/0092-8674(85)90322-8. [DOI] [PubMed] [Google Scholar]
  36. Rennard S. I., Hunninghake G. W., Bitterman P. B., Crystal R. G. Production of fibronectin by the human alveolar macrophage: mechanism for the recruitment of fibroblasts to sites of tissue injury in interstitial lung diseases. Proc Natl Acad Sci U S A. 1981 Nov;78(11):7147–7151. doi: 10.1073/pnas.78.11.7147. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Richter H., Hörmann H. A large cathepsin D-derived fragment from the central part of the fibronectin subunit chains. FEBS Lett. 1983 May 8;155(2):317–320. doi: 10.1016/0014-5793(82)80628-5. [DOI] [PubMed] [Google Scholar]
  38. Seppä H. E., Yamada K. M., Seppä S. T., Silver M. H., Kleinman H. K., Schiffmann E. The cell binding fragment of fibronectin is chemotactic for fibroblasts. Cell Biol Int Rep. 1981 Aug;5(8):813–819. doi: 10.1016/0309-1651(81)90253-8. [DOI] [PubMed] [Google Scholar]
  39. Seppä H., Grotendorst G., Seppä S., Schiffmann E., Martin G. R. Platelet-derived growth factor in chemotactic for fibroblasts. J Cell Biol. 1982 Feb;92(2):584–588. doi: 10.1083/jcb.92.2.584. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Shoshan S. Wound healing. Int Rev Connect Tissue Res. 1981;9:1–26. doi: 10.1016/b978-0-12-363709-3.50007-3. [DOI] [PubMed] [Google Scholar]
  41. Snyderman R., Fudman E. J. Demonstration of a chemotactic factor receptor on macrophages. J Immunol. 1980 Jun;124(6):2754–2757. [PubMed] [Google Scholar]
  42. Tamkun J. W., DeSimone D. W., Fonda D., Patel R. S., Buck C., Horwitz A. F., Hynes R. O. Structure of integrin, a glycoprotein involved in the transmembrane linkage between fibronectin and actin. Cell. 1986 Jul 18;46(2):271–282. doi: 10.1016/0092-8674(86)90744-0. [DOI] [PubMed] [Google Scholar]
  43. Tarone G., Galetto G., Prat M., Comoglio P. M. Cell surface molecules and fibronectin-mediated cell adhesion: effect of proteolytic digestion of membrane proteins. J Cell Biol. 1982 Jul;94(1):179–186. doi: 10.1083/jcb.94.1.179. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Tsukamoto Y., Helsel W. E., Wahl S. M. Macrophage production of fibronectin, a chemoattractant for fibroblasts. J Immunol. 1981 Aug;127(2):673–678. [PubMed] [Google Scholar]
  45. Williams L. T., Snyderman R., Pike M. C., Lefkowitz R. J. Specific receptor sites for chemotactic peptides on human polymorphonuclear leukocytes. Proc Natl Acad Sci U S A. 1977 Mar;74(3):1204–1208. doi: 10.1073/pnas.74.3.1204. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Zigmond S. H., Hirsch J. G. Leukocyte locomotion and chemotaxis. New methods for evaluation, and demonstration of a cell-derived chemotactic factor. J Exp Med. 1973 Feb 1;137(2):387–410. doi: 10.1084/jem.137.2.387. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from The Journal of Cell Biology are provided here courtesy of The Rockefeller University Press

RESOURCES