Skip to main content
PMC home page
The Journal of Cell Biology logoLink to The Journal of Cell Biology
. 1991 Jun 2;113(6):1463–1473. doi: 10.1083/jcb.113.6.1463

Identification of the fibronectin sequences required for assembly of a fibrillar matrix

PMCID: PMC2289042  PMID: 2045422

Abstract

During extracellular matrix assembly, fibronectin (FN) binds to cell surface receptors and initiates fibrillogenesis. As described in this report, matrix assembly has been dissected using recombinant FN polypeptides (recFNs) expressed in mammalian cells via retroviral vectors. RecFNs were assayed for incorporation into the detergent- insoluble cell matrix fraction and for formation of fibrils at the cell surface as detected by indirect immunofluorescence. Biochemical and immunocytochemical data are presented defining the minimum domain requirements for FN fibrillogenesis. The smallest functional recFN is half the size of native FN and contains intact amino- and carboxy- terminal regions with a large internal deletion spanning the collagen binding domain and the first seven type III repeats. Five type I repeats at the amino terminus are required for assembly and have FN binding activity. The dimer structure mediated by the carboxy-terminal interchain disulfide bonds is also essential. Surprisingly, recFNs lacking the RGDS cell binding site formed a significant fibrillar matrix. Therefore, FN-FN interactions and dimeric structure appear to be the major determinants of fibrillogenesis.

Full Text

The Full Text of this article is available as a PDF (2.4 MB).

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., Chen W. T., Yamada K. M. Analysis of fibronectin receptor function with monoclonal antibodies: roles in cell adhesion, migration, matrix assembly, and cytoskeletal organization. J Cell Biol. 1989 Aug;109(2):863–875. doi: 10.1083/jcb.109.2.863. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Allio A. E., McKeown-Longo P. J. Extracellular matrix assembly of cell-derived and plasma-derived fibronectins by substrate-attached fibroblasts. J Cell Physiol. 1988 Jun;135(3):459–466. doi: 10.1002/jcp.1041350313. [DOI] [PubMed] [Google Scholar]
  3. Barry E. L., Mosher D. F. Factor XIIIa-mediated cross-linking of fibronectin in fibroblast cell layers. Cross-linking of cellular and plasma fibronectin and of amino-terminal fibronectin fragments. J Biol Chem. 1989 Mar 5;264(7):4179–4185. [PubMed] [Google Scholar]
  4. Chernousov M. A., Faerman A. I., Frid M. G., Printseva OYu, Koteliansky V. E. Monoclonal antibody to fibronectin which inhibits extracellular matrix assembly. FEBS Lett. 1987 Jun 8;217(1):124–128. doi: 10.1016/0014-5793(87)81255-3. [DOI] [PubMed] [Google Scholar]
  5. Chernousov M. A., Metsis M. L., Koteliansky V. E. Studies of extracellular fibronectin matrix formation with fluoresceinated fibronectin and fibronectin fragments. FEBS Lett. 1985 Apr 22;183(2):365–369. doi: 10.1016/0014-5793(85)80811-5. [DOI] [PubMed] [Google Scholar]
  6. Choi M. G., Hynes R. O. Biosynthesis and processing of fibronectin in NIL.8 hamster cells. J Biol Chem. 1979 Dec 10;254(23):12050–12055. [PubMed] [Google Scholar]
  7. Darribère T., Guida K., Larjava H., Johnson K. E., Yamada K. M., Thiery J. P., Boucaut J. C. In vivo analyses of integrin beta 1 subunit function in fibronectin matrix assembly. J Cell Biol. 1990 May;110(5):1813–1823. doi: 10.1083/jcb.110.5.1813. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Ehrismann R., Chiquet M., Turner D. C. Mode of action of fibronectin in promoting chicken myoblast attachment. Mr = 60,000 gelatin-binding fragment binds native fibronectin. J Biol Chem. 1981 Apr 25;256(8):4056–4062. [PubMed] [Google Scholar]
  9. Ehrismann R., Roth D. E., Eppenberger H. M., Turner D. C. Arrangement of attachment-promoting, self-association, and heparin-binding sites in horse serum fibronectin. J Biol Chem. 1982 Jul 10;257(13):7381–7387. [PubMed] [Google Scholar]
  10. Engvall E., Ruoslahti E. Binding of soluble form of fibroblast surface protein, fibronectin, to collagen. Int J Cancer. 1977 Jul 15;20(1):1–5. doi: 10.1002/ijc.2910200102. [DOI] [PubMed] [Google Scholar]
  11. Fogerty F. J., Akiyama S. K., Yamada K. M., Mosher D. F. Inhibition of binding of fibronectin to matrix assembly sites by anti-integrin (alpha 5 beta 1) antibodies. J Cell Biol. 1990 Aug;111(2):699–708. doi: 10.1083/jcb.111.2.699. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Giancotti F. G., Ruoslahti E. Elevated levels of the alpha 5 beta 1 fibronectin receptor suppress the transformed phenotype of Chinese hamster ovary cells. Cell. 1990 Mar 9;60(5):849–859. doi: 10.1016/0092-8674(90)90098-y. [DOI] [PubMed] [Google Scholar]
  13. Guan J. L., Trevithick J. E., Hynes R. O. Retroviral expression of alternatively spliced forms of rat fibronectin. J Cell Biol. 1990 Mar;110(3):833–847. doi: 10.1083/jcb.110.3.833. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Hayman E. G., Ruoslahti E. Distribution of fetal bovine serum fibronectin and endogenous rat cell fibronectin in extracellular matrix. J Cell Biol. 1979 Oct;83(1):255–259. doi: 10.1083/jcb.83.1.255. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Homandberg G. A., Erickson J. W. Model of fibronectin tertiary structure based on studies of interactions between fragments. Biochemistry. 1986 Nov 4;25(22):6917–6925. doi: 10.1021/bi00370a027. [DOI] [PubMed] [Google Scholar]
  16. Ingham K. C., Brew S. A., Migliorini M. M. Further localization of the gelatin-binding determinants within fibronectin. Active fragments devoid of type II homologous repeat modules. J Biol Chem. 1989 Oct 15;264(29):16977–16980. [PubMed] [Google Scholar]
  17. McDonald J. A. Extracellular matrix assembly. Annu Rev Cell Biol. 1988;4:183–207. doi: 10.1146/annurev.cb.04.110188.001151. [DOI] [PubMed] [Google Scholar]
  18. McDonald J. A., Quade B. J., Broekelmann T. J., LaChance R., Forsman K., Hasegawa E., Akiyama S. Fibronectin's cell-adhesive domain and an amino-terminal matrix assembly domain participate in its assembly into fibroblast pericellular matrix. J Biol Chem. 1987 Mar 5;262(7):2957–2967. [PubMed] [Google Scholar]
  19. McKeown-Longo P. J., Mosher D. F. Binding of plasma fibronectin to cell layers of human skin fibroblasts. J Cell Biol. 1983 Aug;97(2):466–472. doi: 10.1083/jcb.97.2.466. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. McKeown-Longo P. J., Mosher D. F. Interaction of the 70,000-mol-wt amino-terminal fragment of fibronectin with the matrix-assembly receptor of fibroblasts. J Cell Biol. 1985 Feb;100(2):364–374. doi: 10.1083/jcb.100.2.364. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Merril C. R., Goldman D., Van Keuren M. L. Gel protein stains: silver stain. Methods Enzymol. 1984;104:441–447. doi: 10.1016/s0076-6879(84)04111-2. [DOI] [PubMed] [Google Scholar]
  22. Millis A. J., Hoyle M., Mann D. M., Brennan M. J. Incorporation of cellular and plasma fibronectins into smooth muscle cell extracellular matrix in vitro. Proc Natl Acad Sci U S A. 1985 May;82(9):2746–2750. doi: 10.1073/pnas.82.9.2746. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Patel R. S., Odermatt E., Schwarzbauer J. E., Hynes R. O. Organization of the fibronectin gene provides evidence for exon shuffling during evolution. EMBO J. 1987 Sep;6(9):2565–2572. doi: 10.1002/j.1460-2075.1987.tb02545.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Quade B. J., McDonald J. A. Fibronectin's amino-terminal matrix assembly site is located within the 29-kDa amino-terminal domain containing five type I repeats. J Biol Chem. 1988 Dec 25;263(36):19602–19609. [PubMed] [Google Scholar]
  25. Ruoslahti E. Fibronectin and its receptors. Annu Rev Biochem. 1988;57:375–413. doi: 10.1146/annurev.bi.57.070188.002111. [DOI] [PubMed] [Google Scholar]
  26. Schwarzbauer J. E., Mulligan R. C., Hynes R. O. Efficient and stable expression of recombinant fibronectin polypeptides. Proc Natl Acad Sci U S A. 1987 Feb;84(3):754–758. doi: 10.1073/pnas.84.3.754. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Schwarzbauer J. E., Spencer C. S., Wilson C. L. Selective secretion of alternatively spliced fibronectin variants. J Cell Biol. 1989 Dec;109(6 Pt 2):3445–3453. doi: 10.1083/jcb.109.6.3445. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Skorstengaard K., Jensen M. S., Sahl P., Petersen T. E., Magnusson S. Complete primary structure of bovine plasma fibronectin. Eur J Biochem. 1986 Dec 1;161(2):441–453. doi: 10.1111/j.1432-1033.1986.tb10464.x. [DOI] [PubMed] [Google Scholar]
  29. Woods A., Johansson S., Hök M. Fibronectin fibril formation involves cell interactions with two fibronectin domains. Exp Cell Res. 1988 Aug;177(2):272–283. doi: 10.1016/0014-4827(88)90461-2. [DOI] [PubMed] [Google Scholar]
  30. Yamada K. M., Kennedy D. W. Fibroblast cellular and plasma fibronectins are similar but not identical. J Cell Biol. 1979 Feb;80(2):492–498. doi: 10.1083/jcb.80.2.492. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Yamada K. M., Yamada S. S., Pastan I. Cell surface protein partially restores morphology, adhesiveness, and contact inhibition of movement to transformed fibroblasts. Proc Natl Acad Sci U S A. 1976 Apr;73(4):1217–1221. doi: 10.1073/pnas.73.4.1217. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES