Versican is a large chondroitin sulfate proteoglycan produced by several tumor cell types, including malignant melanoma. The expression of increased amounts of versican in the extracellular matrix may play a role in tumor cell growth, adhesion and migration. We have expressed the V3 isoform of versican in human and canine melanoma cell lines. Retroviral overexpression of V3 did not change the morphology of any of the cell lines but markedly reduces cell growth in the V3 versican expressing melanoma cells. The V3-overexpressing melanoma cells retain their diminished growth potential in vivo because primary tumors arising from these cell lines growth more slowly than their vector only counterparts. This effect was accompanied by increases in cell adhesion on hyaluronan and an enhanced ability to migrate on hyaluronan-coated transwell chambers. This enhanced migration is blocked when cells are preincubated with soluble hyaluronan, or anti-CD44 antibodies, suggesting that V3 acts by altering the hyaluronan-CD44 interaction. Hyaluronan content and CD44 expression are not altered in V3-overexpressing cells compared to vector-transduced cells. Our results show that V3 overproduction modulates the in vitro behavior of human and canine melanoma cell lines and reduces their tumorigenicity in vivo.