(A) Western blot analysis of Hs578T, MDA-MB231 and T47D cells stably transduced with either empty MLV back bone (BB) or MLV-HA-RASSF1C (1C) and treated with 1 μg/ml doxycycline for 48 hr. The anti-HA tag antibody detected a HA-RASSF1C fusion protein in cells transduced with 1C but not BB as expected. (B) Hs578T, MDA-MB231, and T47D cells stably transduced with an empty MLV-backbone (BB) or MLV-HA-RASSF1C (1C) vectors were treated with 1 μg/ml doxycycline for 48 hr. Cells were incubated with 3H-thymidine for 6 hr and 3H-thymidine incorporation was assayed as previously described . (C) Hs578T-1C and T47D-1C treated with doxycycline showed an increase in cell proliferation compared to cells not treated with doxycycline, respectively. RASSF1C over-expression did not inhibit but rather caused a small and statistically significant increase in cell proliferation compared to cells stably transduced with empty MLV-backbone vector. Values are mean ± SEM of 8 replicates; experiments were performed at least three independent times.