Axonin-1/TAG-1 Mediates Cell-Cell Adhesion by a Cis-assisted Trans-interaction

Abstract

The neural cell adhesion molecule axonin-1/TAG-1 mediates cell-cell interactions via homophilic and heterophilic contacts. It consists of six Ig and four fibronectin type III domains anchored to the membrane by glycosylphosphatidylinositol. The recently solved crystal structure indicates a module composed of the four N-terminal Ig domains as the contact site between trans-interacting axonin-1 molecules from apposed membranes. Here, we have tested domain-specific monoclonal antibodies for their capacity to interfere with homophilic binding in a cell aggregation assay. The results confirmed the existence of a binding region within the N-terminal Ig domains and identified a second region contributing to homophilic binding on the third and fourth fibronectin domains near the C terminus. The perturbation of each region alone resulted in a complete loss of cell aggregation, suggesting that axonin-1-mediated cell-cell contact results from a cooperative action of two homophilic binding regions. The data support that axonin-1-mediated cell-cell contact is formed by cis-assisted trans-binding. The N-terminal binding regions of axonin-1 establish a linear zipper-like string of trans-interacting axonin-1 molecules alternately provided by the two apposed membranes. The C-terminal binding regions strengthen the cell-cell contact by enhancing the expansion of the linear string into a two-dimensional array via cis-interactions. Cis-assisted trans-binding may be a basic binding mechanism common to many cell adhesion molecules. The neural cell adhesion molecule axonin-1/TAG-1 mediates cell-cell interactions via homophilic and heterophilic contacts. It consists of six Ig and four fibronectin type III domains anchored to the membrane by glycosylphosphatidylinositol. The recently solved crystal structure indicates a module composed of the four N-terminal Ig domains as the contact site between trans-interacting axonin-1 molecules from apposed membranes. Here, we have tested domain-specific monoclonal antibodies for their capacity to interfere with homophilic binding in a cell aggregation assay. The results confirmed the existence of a binding region within the N-terminal Ig domains and identified a second region contributing to homophilic binding on the third and fourth fibronectin domains near the C terminus. The perturbation of each region alone resulted in a complete loss of cell aggregation, suggesting that axonin-1-mediated cell-cell contact results from a cooperative action of two homophilic binding regions. The data support that axonin-1-mediated cell-cell contact is formed by cis-assisted trans-binding. The N-terminal binding regions of axonin-1 establish a linear zipper-like string of trans-interacting axonin-1 molecules alternately provided by the two apposed membranes. The C-terminal binding regions strengthen the cell-cell contact by enhancing the expansion of the linear string into a two-dimensional array via cis-interactions. Cis-assisted trans-binding may be a basic binding mechanism common to many cell adhesion molecules. fibronectin type III monoclonal antibody fluorescein isothiocyanate Interactions between cell adhesion molecules on the surface of neural cells play a key role in several stages of neural development, including cell migration, axon growth and guidance, establishment of synaptic connections, and myelinization. A well characterized representative of a neural cell adhesion molecule is axonin-1 (1Zuellig R.A. Rader C. Schroeder A. Kalousek M.B. von Bohlen und Halbach F. Osterwalder T. Inan C. Stoeckli E.T. Affolter H.-U. Fritz A. Hafen E. Sonderegger P. Eur. J. Biochem. 1992; 204: 453-463Crossref PubMed Scopus (119) Google Scholar), the avian homolog of TAG-1 (2Furley A.J. Morton S.B. Manalo D. Karagogeos D. Dodd J. Jessell T.M. Cell. 1990; 61: 157-170Abstract Full Text PDF PubMed Scopus (472) Google Scholar). Axonin-1 is a member of the immunoglobulin superfamily. It consists of six Ig and four fibronectin type III (Fn)1 domains and is anchored to the cell membrane by a glycosylphosphatidylinositol anchor. Axonin-1 has been found to engage in different interactions with other cell-surface or extracellular matrix components. Axonin-1 on one cell binds homophilically to axonin-1 on another cell (3Rader C. Stoeckli E.T. Ziegler U. Osterwalder T. Kunz B. Sonderegger P. Eur. J. Biochem. 1993; 215: 133-141Crossref PubMed Scopus (76) Google Scholar). Heterophilic interactions of axonin-1 have been reported with the cell adhesion molecules NgCAM (4Kuhn T.B. Stoeckli E.T. Condrau M.A. Rathjen F.G. Sonderegger P. J. Cell Biol. 1991; 115: 1113-1126Crossref PubMed Scopus (248) Google Scholar), NrCAM (5Suter D.M. Pollerberg G.E. Buchstaller A. Giger R.J. Dreyer W.J. Sonderegger P. J. Cell Biol. 1995; 131: 1067-1081Crossref PubMed Scopus (85) Google Scholar), and NCAM (6Milev P. Maurel P. Haring M. Margolis R.K. Margolis R.U. J. Biol. Chem. 1996; 271: 15716-15723Abstract Full Text Full Text PDF PubMed Scopus (187) Google Scholar); the proteoglycans neurocan and phosphacan; and the extracellular matrix component tenascin C (6Milev P. Maurel P. Haring M. Margolis R.K. Margolis R.U. J. Biol. Chem. 1996; 271: 15716-15723Abstract Full Text Full Text PDF PubMed Scopus (187) Google Scholar). As a first step toward a detailed molecular description of the homophilic and heterophilic interactions of axonin-1, we have determined the crystal structure of the first four N-terminal domains of axonin-1 (7Freigang J. Proba K. Leder L. Diederichs K. Sonderegger P. Welte W. Cell. 2000; 101: 425-433Abstract Full Text Full Text PDF PubMed Scopus (139) Google Scholar). We found a U-shaped arrangement of the Ig domains, resulting in the formation of a compact module. In the crystals, these modules are aligned in a string. Adjacent modules exhibit a large edge-to-face contact and are oriented in an antiparallel fashion, with their C termini pointing perpendicularly to the string. This arrangement suggests that cell adhesion by homophilic axonin-1 interactions occurs by the formation of a linear zipper-like array in which the axonin-1 molecules are alternately provided by the two apposed membranes (Fig. 1). In confirmation of this model, mutations in a loop critical for the formation of the zipper result in the loss of the homophilic binding capacity of axonin-1 (7Freigang J. Proba K. Leder L. Diederichs K. Sonderegger P. Welte W. Cell. 2000; 101: 425-433Abstract Full Text Full Text PDF PubMed Scopus (139) Google Scholar). Here, we provide evidence that monoclonal antibodies binding either to the first Ig domain or to the third and fourth Fn domains perturb the formation of aggregates between axonin-1-expressing myeloma cells. Our results indicate that two distinct binding sites, one at the N terminus and one at the C terminus of axonin-1, are involved in axonin-1-mediated cell-cell contact. Perturbing the binding in each region alone resulted in a complete loss of axonin-1-mediated cell-cell contact. Therefore, we propose a model for axonin-1-mediated cell-cell binding based on a cooperative action of the N- and C-terminal binding sites. Axonin-1 was purified from the ocular vitreous fluid of embryonic day 14 chicken embryos as described previously (8Ruegg M.A. Stoeckli E.T. Kuhn T.B. Heller M. Zuellig R. Sonderegger P. EMBO J. 1989; 8: 55-63Crossref PubMed Scopus (66) Google Scholar). For the production of monoclonal antibodies, four 6-week-old BALB/c mice were immunized by subcutaneous injection of 20 or 25 μg of axonin-1 in complete Freund's adjuvant, followed by three to four booster injections of 10–20 μg of axonin-1 in incomplete Freund's adjuvant. 3.5 days before spleens were removed, the mice received a last booster injection of 15 μg of axonin-1 in phosphate-buffered saline. One-half was injected into the tail vein, and the other half into the peritoneum. Monoclonal antibodies were produced according to the protocol given by Fazekas de St. Groth and Scheidegger (9Fazekas de St. Groth S. Scheidegger D. J. Immunol. Meth. 1980; 35: 1-21Crossref PubMed Scopus (1158) Google Scholar). For fusion with spleen cells, the myeloma cell line X63-Ag8.653 was used. All clones resulting from a fusion were tested 11 days after the fusion, and the test was repeated for slowly growing and negative clones at days 13, 16, and 19. Supernatants were tested for their capacity to recognize native purified axonin-1 in a dot immunobinding assay on nitrocellulose. Binding mAbs were visualized with peroxidase-conjugated anti-mouse IgG antibody (Kirkegaard & Perry Laboratories). Positive clones were subcloned at least three times. To test anti-axonin-1 mAbs for the capacity to interfere with the formation of cell aggregates, the axonin-1-expressing myeloma cell line 5u (3Rader C. Stoeckli E.T. Ziegler U. Osterwalder T. Kunz B. Sonderegger P. Eur. J. Biochem. 1993; 215: 133-141Crossref PubMed Scopus (76) Google Scholar) was expanded to an approximate density of 105 cells/ml, centrifuged, and resuspended in fresh medium to a final cell number of 106 cells/ml. For testing aggregation, an aliquot of 50 μl was transferred to each well of a microtiter plate (5 × 104 cells/well). Monoclonal IgG or Fab were added to a final concentration of 50 μg/ml. Polyclonal IgG or Fab were used at 1 mg/ml. To test for aggregation, the cells were incubated at 4 °C for 45 min as described previously (3Rader C. Stoeckli E.T. Ziegler U. Osterwalder T. Kunz B. Sonderegger P. Eur. J. Biochem. 1993; 215: 133-141Crossref PubMed Scopus (76) Google Scholar). Thereafter, cell aggregation was assessed by counting single cells, small aggregates of 5–10 cells, and large aggregates of >10 cells on an inverted phase-contrast microscope. For each measurement, 100 observations were collected and classified into the three groups (single cells, small aggregates of 5–10 cells, and large aggregates of >10 cells). For each condition, the percentage of observations within each category was determined as the between the and the with antibody To test for we used and of axonin-1 were by and into myeloma cells as described previously C. Kunz B. R. Giger R.J. P. P. Sonderegger P. EMBO J. 1996; PubMed Scopus Google Scholar). The axonin-1 were according to the domain that was For in the second Ig domain of axonin-1 was The domain used in this and the that were are the first and last of the in to the of axonin-1 (1Zuellig R.A. Rader C. Schroeder A. Kalousek M.B. von Bohlen und Halbach F. Osterwalder T. Inan C. Stoeckli E.T. Affolter H.-U. Fritz A. Hafen E. Sonderegger P. Eur. J. Biochem. 1992; 204: 453-463Crossref PubMed Scopus (119) Google with the second Ig domain by an first Ig and The between axonin-1 and the cell adhesion molecule were previously described by R. Kunz B. L. R. U. Sonderegger P. Eur. J. Biochem. PubMed Scopus Google Scholar). The of of axonin-1 and of The was composed of of and of myeloma cells were by fusion P. U. S. A. PubMed Scopus Google Scholar) with the To the cells were in medium and in microtiter at °C in clones were by of were the cells were for by axonin-1 or and fluorescein isothiocyanate IgG antibody with were of domain was tested by of the domain was visualized by anti-axonin-1 and IgG antibody To test binding of the myeloma cells or axonin-1 were for at °C with or monoclonal antibody at a concentration of in medium The cells that the antibodies were by with anti-mouse IgG antibody The was with a and with an of axonin-1 was confirmed anti-axonin-1 antibody at a concentration of and IgG antibody at a of of axonin-1 molecules was by a for was to and to an for the was as previously reported for the for the axonin-1 R.J. Ziegler U. Kunz B. Kunz S. Sonderegger P. J. PubMed Scopus Google Scholar). In of the is by the For cells were in The cells were and to in of was and the medium was with The were after To the of axonin-1 and at the cell contact of two cells, we determined the a and was to the between the in the contact of the two cells and the of the of the two cell and To the surface of axonin-1 that are involved in homophilic we tested the capacity of monoclonal antibodies to perturb axonin-1-mediated cell-cell binding and their on the surface of native To antibodies, we used a cell aggregation assay with myeloma cells The axonin-1-expressing myeloma cells large cell aggregates by homophilic interactions between axonin-1 molecules (3Rader C. Stoeckli E.T. Ziegler U. Osterwalder T. Kunz B. Sonderegger P. Eur. J. Biochem. 1993; 215: 133-141Crossref PubMed Scopus (76) Google Scholar). We tested mAbs for their capacity to interfere with myeloma cell In a purified mAbs were tested in their IgG We found mAbs that the formation of with a of these mAbs were tested as Fab for their capacity to axonin-1-expressing myeloma cell aggregates All mAbs the percentage of aggregates composed of >10 cells to or (Fig. aggregates composed of at least cells were their number was of the The of homophilic binding and myeloma cell aggregation In the of these as with anti-axonin-1 Fab antibodies, cell aggregates were was used to that the mAbs that on the formation of myeloma cell aggregates were of binding to cell-surface axonin-1, that their to perturb axonin-1-mediated cell aggregation was to an of their in the of To the of the we used domain of axonin-1 and on the surface of myeloma cells. domains of axonin-1, as by their to the Ig domains or the fibronectin type III domains (1Zuellig R.A. Rader C. Schroeder A. Kalousek M.B. von Bohlen und Halbach F. Osterwalder T. Inan C. Stoeckli E.T. Affolter H.-U. Fritz A. Hafen E. Sonderegger P. Eur. J. Biochem. 1992; 204: 453-463Crossref PubMed Scopus (119) Google Scholar) and by of the axonin-1 R.J. L. Zuellig R.A. Rader C. A. Sonderegger P. Eur. J. Biochem. 1995; PubMed Scopus Google Scholar), were by of axonin-1 were into the axonin-1 (3Rader C. Stoeckli E.T. Ziegler U. Osterwalder T. Kunz B. Sonderegger P. Eur. J. Biochem. 1993; 215: 133-141Crossref PubMed Scopus (76) Google Scholar). these were into myeloma cells by fusion P. U. S. A. PubMed Scopus Google Scholar). that were on the surface of cells. axonin-1 was on myeloma cells. cells axonin-1 and axonin-1 on their surface were used for The of an by a was assessed by anti-mouse IgG antibody and in the on the of binding to myeloma cells a the of a was to a domain of axonin-1 In of three were to Ig domain 1 one to four to one to one to one to Fn domain 1 one to two to and to mAbs to to one of the first three Ig these four were to to the fourth Ig domain C. Kunz B. R. Giger R.J. P. P. Sonderegger P. EMBO J. 1996; PubMed Scopus Google Scholar). mAbs to either the third or fourth Fn of the mAbs that with myeloma cell aggregation and their on the first Ig The other four mAbs and were to on the two C-terminal Fn The of and were on the antibodies of and the of and for The of the 20 antibodies that axonin-1-mediated myeloma cell aggregation were the axonin-1 In we found antibodies with on the domains as the of the antibodies, on the domain and on the third and fourth Fn of the antibodies previously been found to perturb the heterophilic binding of axonin-1 and NgCAM C. Kunz B. R. Giger R.J. P. P. Sonderegger P. EMBO J. 1996; PubMed Scopus Google Scholar). these results indicate that the mAbs that interfere with the homophilic trans-binding of axonin-1 are that are in two of the axonin-1 one at the N terminus and the other at the C terminus. axonin-1 has been found to exhibit a domain arrangement in which the N-terminal domain of the first four Ig domains into with the C-terminal Fn domains C. Kunz B. R. Giger R.J. P. P. Sonderegger P. EMBO J. 1996; PubMed Scopus Google Scholar). In this the binding site for NgCAM is for A. Kunz S. P. Kunz B. Ziegler U. Rader C. Sonderegger P. J. Cell Biol. 1996; PubMed Scopus Google Scholar). axonin-1 by the or Ig domain trans-binding of NgCAM C. Kunz B. R. Giger R.J. P. P. Sonderegger P. EMBO J. 1996; PubMed Scopus Google Scholar). of the N-terminal Ig domains the C-terminal Fn domains is in the axonin-1 the axonin-1 was by a toward the in which the N-terminal Ig domains are for a an structure has been for axonin-1 in homophilic trans-binding (7Freigang J. Proba K. Leder L. Diederichs K. Sonderegger P. Welte W. Cell. 2000; 101: 425-433Abstract Full Text Full Text PDF PubMed Scopus (139) Google Scholar). observations that the site in the Fn region was in the in the of To test the of axonin-1 to with the cell-cell contact was by an structure of axonin-1 as with the we the cell-cell contact regions in of cells different of We axonin-1 in which the Ig domain was C. Kunz B. R. Giger R.J. P. P. Sonderegger P. EMBO J. 1996; PubMed Scopus Google Scholar) as a of axonin-1 with a The cells were with a an of either axonin-1 or the days after the cells were for surface axonin-1 by (Fig. As in the cell contact between axonin-1-expressing cells or at of axonin-1 In cells the of axonin-1 a of axonin-1 the contact with other cells the axonin-1 (Fig. of the cell-surface membranes and cell contact confirmed a of axonin-1 with axonin-1 at cell contact results indicate an of the of axonin-1 to with of cell-cell contact. We found that the of monoclonal antibodies axonin-1 were in two one to the N terminus and the other to the C terminus and Perturbing the binding of each region alone resulted in a complete loss of axonin-1-mediated cell-cell homophilic axonin-1 binding two binding be formed by an of the N-terminal site of with the C-terminal site of with two distinct between and and or a of a between the N termini and a between the C termini of the trans-interacting axonin-1 molecules with the C termini of axonin-1 molecules and The of the interactions of is with and based on The that mAbs either region alone are of axonin-1-mediated cell-cell is for a and For the be that or be to support cell-cell for one that the binding of mAbs to the of axonin-1 perturb a role of the Fn region in the or of the This is by the that axonin-1 has a that is to be by an contact between the N-terminal Ig region and the C-terminal Fn region C. Kunz B. R. Giger R.J. P. P. Sonderegger P. EMBO J. 1996; PubMed Scopus Google Scholar). these we a of two one N-terminal binding and another C-terminal binding of axonin-1 molecules anchored to the membrane have that N- and C-terminal exhibit the capacity of (7Freigang J. Proba K. Leder L. Diederichs K. Sonderegger P. Welte W. Cell. 2000; 101: 425-433Abstract Full Text Full Text PDF PubMed Scopus (139) Google K. J. Karagogeos D. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar). In the of the module formed by the first four Ig domains of axonin-1, (7Freigang J. Proba K. Leder L. Diederichs K. Sonderegger P. Welte W. Cell. 2000; 101: 425-433Abstract Full Text Full Text PDF PubMed Scopus (139) Google Scholar) have identified an edge-to-face contact between with the formation of a linear string of molecules. Adjacent molecules in the string are in antiparallel and their C termini perpendicularly to the of the string. This arrangement a model for axonin-1-mediated cell-cell adhesion in which the N-terminal modules of axonin-1 molecules of apposed membranes in a homophilic by in a zipper-like In support of the model, the of two within the edge-to-face contact region and resulted in a complete loss of axonin-1-mediated cell K. J. Karagogeos D. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar) have reported an aggregation of line cells with the Fn of the of results the existence of a homophilic binding site within the Fn the evidence for this binding site has been in an assay for is that the in in this assay occurs in As previously with axonin-1 and may be in binding to or from the a well T.B. Stoeckli E.T. Condrau M.A. Rathjen F.G. Sonderegger P. J. Cell Biol. 1991; 115: 1113-1126Crossref PubMed Scopus (248) Google A. Kunz S. P. Kunz B. Ziegler U. Rader C. Sonderegger P. J. Cell Biol. 1996; PubMed Scopus Google Scholar). In the of molecules on the surface of cells, a site between Fn in for a cell-cell aggregation via trans-binding is provided to a of the binding from the to the membrane of their to the these binding in the Fn region are to in in between a with axonin-1 and growth of D. Leder L. Sonderegger P. L. J. 133-141Crossref PubMed Scopus Google Scholar) that a between the Fn regions for Therefore, we propose that the binding site in the Fn region mediates between the Fn regions of axonin-1 molecules of the on the results and we propose a cis-assisted trans-binding mechanism for the axonin-1-mediated cell-cell As in the N-terminal binding site on a domain module binds in a linear zipper-like array with molecules alternately provided by the two apposed membranes (7Freigang J. Proba K. Leder L. Diederichs K. Sonderegger P. Welte W. Cell. 2000; 101: 425-433Abstract Full Text Full Text PDF PubMed Scopus (139) Google Scholar). The binding is by the of the C-terminal binding on and which a arrangement of the linear into a two-dimensional This model the observations on interactions between of axonin-1 (7Freigang J. Proba K. Leder L. Diederichs K. Sonderegger P. Welte W. Cell. 2000; 101: 425-433Abstract Full Text Full Text PDF PubMed Scopus (139) Google K. J. Karagogeos D. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar) and the results of antibody perturbation on cell-cell adhesion by The model is in with a of between growth of and a with axonin-1 has been D. Leder L. Sonderegger P. L. J. 133-141Crossref PubMed Scopus Google Scholar). on the of the C termini of axonin-1 modules in the (7Freigang J. Proba K. Leder L. Diederichs K. Sonderegger P. Welte W. Cell. 2000; 101: 425-433Abstract Full Text Full Text PDF PubMed Scopus (139) Google Scholar), the of a array of two Ig domains (7Freigang J. Proba K. Leder L. Diederichs K. Sonderegger P. Welte W. Cell. 2000; 101: 425-433Abstract Full Text Full Text PDF PubMed Scopus (139) Google Scholar), the of a array of two Fn domains A.J. Full Text PDF PubMed Scopus Google Scholar), and the of the and between the Ig and Fn the membranes by axonin-1 in a be at a of that between Fn domains result in a of at least of the domains from the to the membrane or that of Ig and Fn domains are as in PubMed Scopus Google Scholar), a that for an structure of axonin-1 on the of a homophilic between Fn domains a binding model with two is a homophilic between the two Fn domains, membranes be to each other to or with a homophilic between the first Fn domains, an of or be In the structure found by negative of purified axonin-1, the axonin-1 molecule is in a that two in C. Kunz B. R. Giger R.J. P. P. Sonderegger P. EMBO J. 1996; PubMed Scopus Google Scholar). in the of axonin-1, the four N-terminal Ig domains the site S. M. C. Buchstaller A. P. R. Rader C. L. Kunz B. Sonderegger P. J. Cell Biol. PubMed Scopus Google Scholar) are to the cell on this the of heterophilic trans-binding between axonin-1 and NgCAM on the surface of different myeloma cells A. Kunz S. P. Kunz B. Ziegler U. Rader C. Sonderegger P. J. Cell Biol. 1996; PubMed Scopus Google Scholar) is the of axonin-1, in which the Ig domain was a trans-binding of NgCAM C. Kunz B. R. Giger R.J. P. P. Sonderegger P. EMBO J. 1996; PubMed Scopus Google Scholar). It is for that the of an Ig domain in the of the molecule the and the structure in which the N-terminal module is for a trans-binding In this we have found that the of axonin-1 an to with the cell contact as by the of an of the at cell contact in cells (Fig. As is the of trans-binding to the structure of the may the establishment of homophilic trans-binding by the from the to the The that a is for the establishment of homophilic binding is by the between cell-cell and binding Axonin-1 on the surface of myeloma cells in the mediates cell aggregation by homophilic binding C. Stoeckli E.T. Ziegler U. Osterwalder T. Kunz B. Sonderegger P. Eur. J. Biochem. 1993; 215: 133-141Crossref PubMed Scopus (76) Google Scholar), homophilic binding is axonin-1 is to the surface of via a with on the a between the of the cell and the has been for NgCAM (4Kuhn T.B. Stoeckli E.T. Condrau M.A. Rathjen F.G. Sonderegger P. J. Cell Biol. 1991; 115: 1113-1126Crossref PubMed Scopus (248) Google A. Kunz S. P. Kunz B. Ziegler U. Rader C. Sonderegger P. J. Cell Biol. 1996; PubMed Scopus Google Scholar) and NrCAM (5Suter D.M. Pollerberg G.E. Buchstaller A. Giger R.J. Dreyer W.J. Sonderegger P. J. Cell Biol. 1995; 131: 1067-1081Crossref PubMed Scopus (85) Google D. Stoeckli E.T. Kunz S. K. Rader C. Kunz B. Buchstaller A. D.M. Dreyer W.J. Sonderegger P. J. Cell Biol. 2000; PubMed Scopus Google Scholar). As a one may that binding to the via groups on the surface of each axonin-1 the of axonin-1 that that may be of the that is for an in homophilic In these results indicate that the from a to an of axonin-1 is for the establishment of homophilic trans-binding between axonin-1 molecules of different cells. The Fn by binding to the and the of the module for trans-binding. Therefore, the of a binding to Fn be an the into the structure and with the of the Fn region by antibodies trans-binding. The loss of cell-cell binding in the of mAbs Fn be by a Therefore, we that the from the to the the of the modules for homophilic in with another In of the homophilic binding site in Fn K. J. Karagogeos D. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar), an with the of axonin-1 molecules to the for the of the mAbs A of two binding sites, the one in and the other in has been found for cell-cell interactions by a D. J. Biol. PubMed Scopus Google Scholar). In the site is within the N-terminal domain and by has been found to have an role for the establishment of trans-binding. In the cis-assisted trans-binding model for axonin-1-mediated cell-cell as we that is for the establishment of trans-binding in the of axonin-1 the linear of modules formed in the of the Fn the Fn regions of axonin-1 a role after the trans-binding by the modules has been It the expansion and of the cell-cell contact by the formation of linear of the first of Fn domains of axonin-1 of two linear has axonin-1 molecules of the one string in with axonin-1 molecules of the other resulting in the of the between Fn domains be as single be and an a arrangement of is A of the of axonin-1 molecules be by the of axonin-1 molecules from the contact The of 1 and was by