Differential Effects of Protein Kinase A on Ras Effector Pathways

ABSTRACT

Ras mutants with the ability to interact with different effectors have played a critical role in the identification of Ras-dependent signaling pathways. We used two mutants, Ras^S35 and Ras^G37, which differ in their ability to bind Raf-1, to examine Ras-dependent signaling in thyroid epithelial cells. Wistar rat thyroid cells are dependent upon thyrotropin (TSH) for growth. Although TSH-stimulated mitogenesis requires Ras, TSH activates protein kinase A (PKA) and downregulates signaling through Raf and the mitogen-activated protein kinase (MAPK) cascade. Cells expressing Ras^S35, a mutant which binds Raf, or Ras^G37, a mutant which binds RalGDS, exhibited TSH-independent proliferation. Ras^S35stimulated morphological transformation and anchorage-independent growth. Ras^G37 stimulated proliferation but not transformation as measured by these indices. TSH exerted markedly different effects on the Ras mutants and transiently repressed MAPK phosphorylation in Ras^S35-expressing cells. In contrast, TSH stimulated MAPK phosphorylation and growth in cells expressing Ras^G37. The Ras mutants, in turn, exerted differential effects on TSH signaling. Ras^S35 abolished TSH-stimulated changes in cell morphology and thyroglobulin expression, while Ras^G37 had no effect on these activities. Together, the data indicate that cross talk between Ras and PKA discriminates between distinct Ras effector pathways.

ACKNOWLEDGMENTS

We thank Lisa Cass for helpful discussions and Jonathan Nissanov for technical advice.

This work was supported by PHS grants DK45696 and DK 02494 awarded to J.M.