HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Submit a Letter to Editor Alert me when this article is cited Alert me when eLetters are posted Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Liao, Y.-F. Articles by De Water, L. V. Search for Related Content PubMed PubMed Citation Articles by Liao, Y.-F. Articles by De Water, L. V. Social Bookmarking                      What's this?

J Biol Chem, Vol. 274, Issue 25, 17876-17884, June 18, 1999

Identification of Two Amino Acids within the EIIIA (ED-A) Segment of Fibronectin Constituting the Epitope for Two Function-blocking Monoclonal Antibodies

From the Center for Engineering in Medicine and Surgical Services, Massachusetts General Hospital and Harvard Medical School, the Shriners Burns Hospital, Boston, Massachusetts 02114

Alternative splicing of the fibronectin gene transcript gives rise to a group of adhesive glycoproteins showing restricted spatial and temporal expression during embryonic development, tumor growth, and tissue repair. Alternative splicing occurs in three segments termed EIIIB, EIIIA, and V. The EIIIA (or ED-A) segment of fibronectin is expressed prominently but transiently in healing wounds coincident with fibroblast expression of an activation marker, smooth muscle cell -actin. A monoclonal antibody (IST-9) to the EIIIA segment blocks transforming growth factor--mediated smooth muscle cell -actin expression by fibroblasts in culture. A second monoclonal antibody (DH1) blocks chondrocyte condensation in chicken embryos. We find that IST-9 and DH1 react with human, rat, and chicken but not with mouse or frog EIIIA, suggesting that His⁴⁴ may be important for antibody binding. A series of deletion mutants of rat EIIIA, constructed as glutathione S-transferase fusion proteins, do not react with either IST-9, DH1, or a third monoclonal antibody (3E2). Mutations of pairs of amino acids to alanine have little effect, except for either (Val³⁴Thr³⁵) or (Tyr³⁶Ser³⁷), which are located in a  strand upstream from His⁴⁴. For these double mutants, the binding to all three monoclonal antibodies is markedly reduced. By contrast, single mutants at Thr³⁵, Tyr³⁶, or Ser³⁷ retain full activity, suggesting that the epitope for these antibodies is determined in part by conformation. Alanine-scanning mutagenesis of rat EIIIA demonstrates the importance of Ile⁴³ and His⁴⁴ for binding. Mutation of frog EIIIA (normally Val⁴³Lys⁴⁴) to rat (Ile⁴³His⁴⁴) is sufficient to restore fully IST-9 binding and much of the activity of DH1 and 3E2. Our findings demonstrate that the function-blocking antibodies, IST-9 and DH1, bind to the Ile⁴³ and His⁴⁴ residues in a conformationally dependent fashion, implicating the loop region encompassing both residues as critical for mediating EIIIA function.

CiteULike

Complore

Connotea

Del.icio.us

Digg

Reddit

Technorati

This article has been cited by other articles:

				A. V. Shinde, C. Bystroff, C. Wang, M. G. Vogelezang, P. A. Vincent, R. O. Hynes, and L. Van De Water Identification of the Peptide Sequences within the EIIIA (EDA) Segment of Fibronectin That Mediate Integrin {alpha}9{beta}1-dependent Cellular Activities J. Biol. Chem., February 1, 2008; 283(5): 2858 - 2870. [Abstract] [Full Text] [PDF]

				F. Poobalarahi, C. F. Baicu, and A. D. Bradshaw Cardiac myofibroblasts differentiated in 3D culture exhibit distinct changes in collagen I production, processing, and matrix deposition Am J Physiol Heart Circ Physiol, December 1, 2006; 291(6): H2924 - H2932. [Abstract] [Full Text] [PDF]

				R. M. Klein, M. Zheng, A. Ambesi, L. Van De Water, and P. J. McKeown-Longo Stimulation of extracellular matrix remodeling by the first type III repeat in fibronectin J. Cell Sci., November 15, 2003; 116(22): 4663 - 4674. [Abstract] [Full Text] [PDF]

				Y.-F. Liao, P. J. Gotwals, V. E. Koteliansky, D. Sheppard, and L. Van De Water The EIIIA Segment of Fibronectin Is a Ligand for Integrins alpha 9beta 1 and alpha 4beta 1 Providing a Novel Mechanism for Regulating Cell Adhesion by Alternative Splicing J. Biol. Chem., April 19, 2002; 277(17): 14467 - 14474. [Abstract] [Full Text] [PDF]

				V. Dugina, L. Fontao, C. Chaponnier, J. Vasiliev, and G. Gabbiani Focal adhesion features during myofibroblastic differentiation are controlled by intracellular and extracellular factors J. Cell Sci., March 11, 2002; 114(18): 3285 - 3296. [Abstract] [Full Text] [PDF]

				B. Hinz, D. Mastrangelo, C. E. Iselin, C. Chaponnier, and G. Gabbiani Mechanical Tension Controls Granulation Tissue Contractile Activity and Myofibroblast Differentiation Am. J. Pathol., September 1, 2001; 159(3): 1009 - 1020. [Abstract] [Full Text] [PDF]

				S. D.J.M. Kanters, J.-D. Banga, A. Algra, R. C.J.M. Frijns, J. J. Beutler, and R. Fijnheer Plasma Levels of Cellular Fibronectin in Diabetes Diabetes Care, February 1, 2001; 24(2): 323 - 327. [Abstract] [Full Text]