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A synthetic peptide from the heparin-binding domain III (repeats III4-5) of fibronectin promotes stress fiber and focal adhesion formation in melanoma cells

AuthorsMoyano, José V.; Maqueda, Alfredo; Albar, Juan Pablo; García-Pardo, Angeles
KeywordsCell-surface proteoglycans
cytoskeletal reorganization
fibronectin heparin-binding domain III (Hep III) domain
heparin-binding peptide
Issue DateJan-2003
PublisherPortland Press
CitationBiochemical Journal, 371 (2) : 565-571 (2003)
AbstractCell adhesion to fibronectin results in formation of actin stress fibres and focal adhesions. In fibroblasts, this response requires two co-operative signals provided by interactions of the RGD sequence with a5b1 integrin and the heparin-binding domain II (Hep II) domain with syndecan-4. Within Hep II, this activity was mapped to repeat III13 and to the peptide FN-C/H-V(WQPPRARITGY, repeat III14). We previously described that the synthetic heparin-binding peptide/III5 (HBP/III5) (WTPPRAQITGYRLTVGLTRR, repeat III5) binds heparin and mediates cell adhesion via chondroitin sulphate proteoglycans. We have now studied whether HBP/III5 co-operates with a5b1 and drives a full cytoskeletal response in melanoma cells. SKMEL-178 cells attached and spread on the RGD-containing FNIII7–FNIII10 (FNIII7–10) fragment, but did not form stress fibres or focal adhesions. Co-immobilization of HBP/III5 with FNIII7–10 or adding soluble HBP/III5 to cells prespread on FNIII7–10, effectively induced these structures. Cell transfection with dominant-negative N19RhoA, a member of the small GTPase family, abolished the HBP/III5 effect. Both chondroitinase and heparitinase diminished focal adhesions, indicating that both types of proteoglycans bound HBP/III5 in melanoma cells. We have mapped the active sequence of HBP/III5 to YRLTVGLTRR, which is a novel sequence in fibronectin with focal-adhesion-promoting activity. The last two arginine (R) residues of this sequence are required for activity, since their replacement by alanine completely abrogated the HBP/III5 cytoskeletal effect. Moreover, this sequence is also active in the context of large fibronectin fragments. Our results establish that the Hep III region provides co-operative signals to a5b1 for the progression of the cytoskeletal response and that these include activation of RhoA
Description7 páginas, 3 tablas -- PAGS nros. 565-571
Publisher version (URL)http:dx.doi.org/10.1042/BJ20021344
Appears in Collections:(CIB) Artículos
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