| STAT3 is a latent transcription factor activated by different stimuli, including cytokines, growth factors and oncogenes. The biological responses of STAT3 include cell proliferation, survival and differentiation and these different effects are determined by the stimulus and cell context.; To overcome the embryonic lethality of STAT3 deletion in mice, our lab has developed conditional STAT3 knockout mice using the Cre-LoxP system. We generated primary fibroblasts from STAT3 “floxed” mice, immortalized them and introduced Cre recombinase to generate STAT3 null cells. STAT3 WT and KO cells were then assayed for different aspects of growth factor responses, as well as for transformation by H-RasV12 and v-Src. Proliferation, survival and growth-factor responses were largely unaffected by the absence of STAT3, except for a significant decrease in c-fos expression following PDGF stimulation, which we found was STAT3 and Src dependent.; When H-RasV12 or v-Src was expressed in STAT3 null cells, transformation was greatly attenuated, as measured by growth in soft agar as well as tumor formation in nude mice. However, the mechanism by which STAT3 was required for H-RasV12 and v-Src transformation was fundamentally different. In the case of v-Src, STAT3 was phosphorylated by v-Src, leading to activation of its transcriptional function. Specifically, STAT3 was required for mediating loss-of-adhesion induced by v-Src through activation of Focal Adhesion Kinase. In the case of H-RasV12 STAT3 was not phosphorylated or activated, and a transcriptionally incompetent STAT3 was able to rescue transformation by H-RasV12. We showed that STAT3 was required for activation of RaIA, which is required for H-RasV12 transformation, specifically for metastasis formation. We confirmed the requirement for STAT3 by metastasis assays in mice, showing that STAT3 KO cells were unable to form metastases in mice, while cells reconstituted with transcriptionally incompetent STAT3 formed metastases at high efficiency, correlating with increased Ral activity.; In summary, we identified multiple roles for STAT3 in normal cell functions and oncogenesis. We showed that STAT3 is required for transformation by two major yet distinct oncogenes, and plays a fundamentally different role in each case. We propose a novel mechanism for STAT3 function that is independent of its transcriptional activation. |
