The in vitro and in vivo effects of several Wnt family members have been studied using stably transfected HC11 cells, a clonal mammary epithelial cell line derived from a midpregnant mouse mammary gland capable of hormone-dependent differentiation in vitro. Differential effects of Wnt-1, Wnt-2, and Wnt-7B expression were observed both on the morphology of confluent HC11 cells and on the pattern of E-cadherin expression. Wnt-7B had no apparent effect on HC11 cell morphology or E-cadherin expression, as compared to mock-transfected HC11 cells. Injection of stably transfected pools of Wnt-1, Wnt-2, Wnt-7B, and mock-transfected cells into the cleared fat pad of syngeneic BALB/c mice generated reproducible outgrowths after 8 or 12 weeks. Mock-transfected cells produced outgrowths that exhibited some morphologically normal ductal and alveolar-like structures. However, no morphologically normal structures were observed in the fat pads containing Wnt-transfected cells. Instead, these outgrowths were characterized by significant fibrosis, epithelial hyperplasia, and multiple sites of growth. In contrast to the lack of an observed effect in vitro, palpable adenocarcinomas were observed 12 weeks after injection of the Wnt-7B-transfected HC11 cells. These tumors contained significant regions of hyperplastic and transformed epithelium and lacked the fibrotic phenotype observed in the Wnt-1 and -2 outgrowths. These results support the hypothesis that different Wnt family members may elicit distinct functional effects and reinforce the need to perform simultaneous comparisons of Wnt function both in vitro and in vivo. Stably transfected HC11 cells provide a useful model system in which to elucidate the function of different Wnt family members.