The Role of Phosphatidylinositol 3-Kinase, Rho Family GTPases, and STAT3 in Ros-induced Cell Transformation

Mai-Thao Le,Kevin T. Nguyen,Shailaja Uttamsingh,Pallavi Sachdev,Joseph L.-K. Chan,Cong S. Zong,Lu-Hai Wang,Monica Bhanot

doi:10.1074/jbc.m108166200

Abstract

Using loss-of-function mutants of Ros and inducible epidermal growth factor receptor-Ros chimeras we investigated the role of various signaling pathways in Ros-induced cell transformation. Inhibition of the mitogen-activated protein kinase (MAPK) pathway with the MEK (MAP/extracellular signal-regulated kinase kinase) inhibitor PD98059 had little effect on the Ros-induced monolayer and anchorage-independent growth of chicken embryo fibroblasts and NIH3T3 cells even though more than 70% of the MAPK was inhibited. In contrast, inhibiting the phosphatidylinositol 3-kinase (PI3K) pathway with the drug LY294002, a dominant negative mutant of PI3K, Deltap85, or the phosphatidylinositol phosphatase PTEN (phosphatase and tensin homologue deleted in chromosome ten) resulted in a dramatic reduction of v-Ros- and epidermal growth factor receptor-Ros-promoted anchorage-independent growth of chicken embryo fibroblasts and NIH3T3 cells, respectively. Parallel and downstream components of PI3K signaling such as the Rho family GTPases (Rac, Rho, Cdc42) and the survival factor Akt were all shown to contribute to Ros-induced anchorage-independent growth, although Rac appeared to be less important for Ros-induced colony formation in NIH3T3 cells. Furthermore, the transformation-attenuated v-Ros mutants F419 and DI could be complemented by constitutively active mutants of PI3K and Akt. Finally, we found that overexpressing a constitutively active mutant of STAT3 (STAT3C) conferred a resistance to the inhibition of Ros-induced anchorage-independent growth by LY294002, suggesting a possible overlap of functions between PI3K and STAT3 signaling in mediating Ros-induced anchorage-independent growth.

Highlights

The proto-oncogene c-Ros encodes a receptor protein-tyrosine kinase (RPT kinase)1 that shares structural homology with the
The drug inhibitors that were utilized included PD98059, which is a specific inhibitor of MEK in the mitogenactivated protein kinase (MAPK) pathway, LY294002, which is a specific inhibitor of phosphatidylinositol 3-kinase (PI3K), and rapamycin, which is a specific inhibitor of mammalian TOR (mTOR)/FRAP that is required to activate p70S6K
We have shown that activation of the EGFR-Ros chimeric receptor overexpressed in NIH3T3 cells results in activation of MAPK, PI3K, STAT3, and Vav3 and leads to transformation of the cells [8, 9, 34]

Summary

Introduction

The proto-oncogene c-Ros encodes a receptor protein-tyrosine kinase (RPT kinase)1 that shares structural homology with the. Effect of PD98059, LY294002, and Rapamycin on the Monolayer Growth of Normal and Ros-expressing CEF and NIH3T3 Cells—The MAPK, PI3K, and p70S6K signaling pathways have been associated with growth factor receptor-mediated mitogenicity in various cell lines.

Results

Conclusion