Controlling Ras activity by manipulating its localization

Abstract

Proteins of the Ras family of small GTPases play a major role in the transduction of extracellular signals, especially growth factor signals, into the cell. Ras proteins thereby participate in the regulation of critical processes like cellular growth and proliferation. Due to this central role, gain-of-function mutations of these proteins can have severe consequences for the mutated cells including uncontrolled proliferation and these mutations are often connected to development of diseases including cancers of various tissues. Thereby, plasma membrane localization is crucial for the activity of Ras proteins. They not just recruit their effectors to the plasma membrane to facilitate their activation, the activation of Ras proteins is also highly dependent on plasma membrane localization. Due to the dependence of Ras activity on plasma membrane localization, modified Ras proteins, whose localization within the cell can be controlled externally, could be useful tools to control Ras activity. Hence, in this work plasma membrane recruitable Ras proteins have been engineered that incorporate two different dimerization approaches based on chemically induced dimerization allowing translocation of either wild type or oncogenically mutated KRas proteins from the cytosol to the plasma membrane in a highly controllable manner. Contrary to the widespread believe that Ras carrying an oncogenic mutation is always in the GTP-bound active state, cytosolic recruitable Ras is just partially active. However, the inactive fraction of recruitable oncogenic Ras becomes activated upon plasma membrane recruitment suggesting that RasGEFs are necessary to maintain the activity of oncogenic Ras at the plasma membrane. Further, recruitment of oncogenic Ras to the plasma membrane sensitized the cellular ERK phosphorylation response to growth factor stimulation. Using light-induced recruitment of oncogenic Ras to the plasma membrane, it could be shown that Ras activity can be propagated to surrounding cells highlighting a potential mechanism allowing Ras transformed cells to influence adjacent cells.