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Adhesive functions in fibronectin's alternatively-spliced ED(a) segment
| Title: | Adhesive functions in fibronectin's alternatively-spliced ED(a) segment |
| Author: | Xia, Ping |
| Description: | EDa (EIIIA) is an alternatively-spliced type III homology repeat present in cellular fibronectins (cFNs) but not in plasma fibronectin (pFN). Potential adhesion-promoting activity was investigated for EDa and its two neighboring repeats III11 and III12. Recombinant protein corresponding to EDa alone promoted Balb/c 3T3 cell attachment, as did neighboring repeats III11 and III12 when tested as single repeats. Activities were even higher for recombinant proteins containing two or three repeats. While EDa alone exhibited 40-60% of the attachment activity of human pFN depending upon cell type, EDa with both neighboring repeats displayed 70-90% of pFN activity. Besides attachment, the recombinant substrata also supported limited cell spreading. 3T3 cells on the EDa substratum displayed round cell bodies with filopodia contacting the substratum. Cells on III11-EDa-III12 displayed spindle-shaped cell bodies and pseudopods in contact with the substratum. The two-repeat molecule of III11-III12 gave an intermediate response. Moreover, adhesion activities towards these recombinant proteins were oncogene-regulated: comple tely abrogated by two different ras oncogenes, unaffected by the sis oncogene, and elevated by the src oncogene aFunctional complementarity was noted between EDa recombinant molecule and pFN. Co-coating substrata with EDa and a suboptimal concentration of pFN led to increased attachment and extensive spreading of v-src-transformed 3T3 cells relative to that found on substrata of suboptimal pFN or EDa alone. This complementarity requires as little as 1 μg/ml EDa in the adsorbing mixture and displays sequence specificity for only EDa (i.e., III11 or III12 was without effect.). Furthermore, stress fibers and focal contacts were inducible on the EDa:PFN mixture, suggesting the full adhesion-promoting competence of the heterologous substrata aSynergy in promoting post-attachment adhesion responses among III11, EDa and III12 was also observed. This conclusion is supported by two lines of evidence: (a) spreading on two-repeat molecules was better than spreading on single-repeat molecules, and (b) only the three-repeat molecule was competent for inducing stress fibers and focal adhesions. Experimental evidence indicated that glycosaminoglycans did not participate in these adhesion processes; neither did β1-containing integrins. Affinity chromatography was employed to isolate the putative receptor(s) to no avail. These studies reveal a new adhesion promoting activity in fibronectin molecules whose function is more effectively presented by cFNs than pFN, which may indicate the advantage of cFNs in various physiological and pathological conditions where expression of EDa sequence is elevated. |
| Permanent Link: |
http://rave.ohiolink.edu/etdc/view?acc_num=case1057591352
http://hdl.handle.net/2374.OX/16202 |
| Date: | 1996 |
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