In murine and rat fibroblasts, activation of several G proteins (Gi2, Gq, G12) can stimulate cell growth or transformation, and can induce tumor formation in nude mice; contrastingly, inactivation of Gi2 inhibits fibroblast proliferation in vitro. We investigated whether it is possible to modulate malignant cell growth in vitro and in vivo through alteration of Gi2 protein function. To do so, we introduced mutated alpha subunits of Gi2 (alpha i2) in CL19 cells, a clone of the murine melanoma cell line K-1735. When we did this, a constitutively activated mutant (alpha i2-Q205L) and a dominant negative mutant (alpha i2-G204A) of alpha i2 were stably expressed in CL19 cells. We found that the in vitro motility of all alpha i2-transfected CL19 cells was increased; however, overexpression and alteration of the function of Gi2 did not increase metastasis formation by CL19 cells in nude mice. Expression of alpha i2-Q205L conferred a limited growth advantage to CL19 cells in vitro; in vivo, tumor formation and size, and overall survival of animals injected with CL19 cells expressing alpha i2-Q205L, were similar to controls. In contrast, expression of the inactive alpha i2-G204A mutant inhibited CL19 growth in vitro by at least 50% in all conditions tested, and mice injected with cells expressing the alpha i2-G204A mutant showed delayed tumor formation, reduced tumor size, and longer survival. We conclude that Gi2 proteins contribute to malignant cell growth, and more importantly, that inactivation of Gi2 proteins can inhibit proliferation of melanoma cells and possibly of other malignant cells both in vitro and in vivo.