Breast and prostatic carcinomas, melanoma, and endothelial cell lines are chemoattracted by medium conditioned by mature osteoblasts. The chemoattractant for endothelial cells was identified with C3, carboxyl-terminal trimer of pro-collagen type I. We report that C3 induces directional migration and proliferation, the expression of tissue inhibitor of metalloproteinases-2, prometalloproteinase-2 and -9, and their activation in MDA MB231 cells, without changing the expression of tissue inhibitor of metalloproteinases-1 and of metalloproteinase-14. Antiserum against metalloproteinase-2 or -9 or -14, tissue inhibitor of metalloproteinases-1, or GM6001 inhibits the C3-induced migration. Urokinase and its receptor are detected and unchanged upon exposure to C3. The antibody against urokinase or addition of plasminogen activator inhibitor inhibits migration. Blocking antibodies to integrins ␣ 2 , ␣ 6 ,  1 , and  3 inhibit chemotaxis and do not change urokinase and urokinase receptor expression. Blockage of ␣ 2 ,  1 , and  3 integrins affect differently the induction by C3 of pro-metalloproteinase-2 and -9 and of tissue inhibitor of metalloproteinases-2. Chemotaxis to C3 is also inhibited by genistein, by pertussis toxin, which also inhibits C3-induced pro-metalloproteinase -2 and -9, but not urokinase expression. Wortmannin partially inhibits C3-induced cell migration. Other, but not all, breast carcinoma lines tested responded to C3 with migration and pro-metalloproteinase-2 induction. Presently C3 is the only agent known to induce migration specifically of both endothelial and breast carcinoma cells. The mitogenic and motogenic role of C3 in vitro might prefigure a role in in vivo carcinogenesis and in the establishment of metastasis.Mature osteoblasts conditioned medium (CM), 1 collected when sustained biosynthesis of collagen type I occurs during osteogenesis in vitro from differentiating cultures of rat, rabbit, human osteoblasts, and conditionally transformed human osteoblast-like line HFO, induces directional migration of endothelial and melanoma and breast and prostatic carcinoma cells but not of normal cells or of transformed mesenchymal cells in the classical in vitro double chamber Boyden assay (1-4).The molecule chemoattractant for endothelial cells was purified from rat osteoblast CM, and it identifies by sequence, immunoreactivity, and molecular size with the trimer carboxylterminal of type I pro-collagen, C3 (4). C3 is a by-product of the processing of pro-collagen by pro-collagenase C (or BMP-1), preliminary to the formation of collagen fibrils (5).We investigated whether MDA MB231 breast carcinoma cells, which are stimulated by mature osteoblasts CM to directional migration, are induced by C3 to activate migratory and proliferation responses. It is recognized that the stroma of the tumors plays instructive and active roles toward tumor cells growth and invasiveness and that tumor-associated stromal fibroblasts undergo changes in the biosynthetic phenotype (6), assume the production of factors supp...