Cell surface molecules that bind Fibronectin's matrix assembly domain

Andrew Harold Limper, Bradley J. Quade, Rhonda M. Lachance, Thomas M. Birkenmeier, Tasneem S. Rangwala, John A. McDonald

Research output: Contribution to journalArticle

26 Citations (Scopus)

Abstract

The assembly of fibronectin into disulfide crosslinked extracellular matrices requires the interaction of mesenchymal cells with two distinct sites on fibronectin, the Arg-Gly-Asp cell adhesive site and an amino-terminal site contained within the first five type I homologous repeats (Quade, B. J., and McDonald, J. A. (1988) J. Biol. Chem. 263, 19602-19609). Proteolytically derived 29-kDa fragments of fibronectin (29kDa) containing these repeats bind to monolayers of cultured fibroblasts and inhibit fibronectin matrix assembly. The cell surface molecules interacting with fibronectin's 29-kDa matrix assembly domain have resisted purification using conventional methods such as affinity chromatography. Accordingly, in order to identify molecules which bind this fragment, 125I-labeled 29kDa was allowed to bind to fibroblast monolayers and chemically cross-linked to the cell surface with bis(sulfosuccinimidyl) suberate. Extraction of the cross-linked cell layer yielded radiolabeled complexes of 56, 150, and 280 kDa. Formation of these crosslinked complexes was specifically inhibited by the addition of excess unlabeled 29kDa but was unaffected by the presence of fibronectin fragments containing other type I repeats outside of the 29kDa matrix assembly domain. The cross-linked complexes were insoluble in nondenaturing detergents but soluble when denatured and reduced, suggesting that 29kDa may be cross-linked to components of the pericellular matrix. Immunoprecipitation of cross-linked cell extracts with a polyclonal antibody to fibronectin that does not recognize the amino terminus demonstrate that the 280-kDa band contains 29kDa cross-linked to fibronectin present on the cell surface. Formation of the 150-kDa complex was inhibited by EDTA, suggesting that divalent cations are required for its formation. Although the molecular mass and divalent cation requirement suggest that the 150-kDa complex may be related to an integrin, this complex was not immunoprecipitated by polyclonal antibodies generated to the α5β1 integrin fibronectin receptor.

Original languageEnglish (US)
Pages (from-to)9697-9702
Number of pages6
JournalJournal of Biological Chemistry
Volume266
Issue number15
StatePublished - 1991
Externally publishedYes

Fingerprint

Fibronectins
Molecules
Divalent Cations
Fibroblasts
Integrins
Monolayers
Integrin alpha5beta1
Affinity chromatography
Antibodies
Molecular mass
Cell Extracts
Affinity Chromatography
Immunoprecipitation
Edetic Acid
Cell Communication
Disulfides
Adhesives
Detergents
Purification
Extracellular Matrix

ASJC Scopus subject areas

  • Biochemistry

Cite this

Limper, A. H., Quade, B. J., Lachance, R. M., Birkenmeier, T. M., Rangwala, T. S., & McDonald, J. A. (1991). Cell surface molecules that bind Fibronectin's matrix assembly domain. Journal of Biological Chemistry, 266(15), 9697-9702.

Cell surface molecules that bind Fibronectin's matrix assembly domain. / Limper, Andrew Harold; Quade, Bradley J.; Lachance, Rhonda M.; Birkenmeier, Thomas M.; Rangwala, Tasneem S.; McDonald, John A.

In: Journal of Biological Chemistry, Vol. 266, No. 15, 1991, p. 9697-9702.

Research output: Contribution to journalArticle

Limper, AH, Quade, BJ, Lachance, RM, Birkenmeier, TM, Rangwala, TS & McDonald, JA 1991, 'Cell surface molecules that bind Fibronectin's matrix assembly domain', Journal of Biological Chemistry, vol. 266, no. 15, pp. 9697-9702.
Limper AH, Quade BJ, Lachance RM, Birkenmeier TM, Rangwala TS, McDonald JA. Cell surface molecules that bind Fibronectin's matrix assembly domain. Journal of Biological Chemistry. 1991;266(15):9697-9702.
Limper, Andrew Harold ; Quade, Bradley J. ; Lachance, Rhonda M. ; Birkenmeier, Thomas M. ; Rangwala, Tasneem S. ; McDonald, John A. / Cell surface molecules that bind Fibronectin's matrix assembly domain. In: Journal of Biological Chemistry. 1991 ; Vol. 266, No. 15. pp. 9697-9702.
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