The role of phosphorylation in the control of Ras activity and localisation in S. cerevisiae

Ras proteins are small GTPases that act as molecular switches within cells that link extracellular stimuli to intracellular effectors. Ras proteins play a conserved role in the control of both cell growth and proliferation. As a result, mutations that induce the constitutive activation of Ras proteins are often associated with changes in cell behaviour that can lead to disease, such as human cancer. The localisation of Ras is crucial for its function and this is controlled by post-translational modifications. However, the roles for such modifications in regulating Ras localisation and its activity are poorly understood. We have identified that the phosphorylation of Serine225 of Ras2, a protein that is essential for the control of both growth and proliferation in S. cerevisiae, plays an important role in the regulation of its localisation and activity. Modification of this residue leads to changes in the distribution of GTP-bound Ras2 within the cell. This drives cells towards a novel state of growth cessation that is dependent upon the activity of the cAMP/PKA signalling pathway. We show that this quiescent state is characterised by an uncoupling of cytoplasmic and nuclear process that govern cell growth and division. We suggest that cells can escape growth arrest and re-engage in the cell cycle if the Ras/cAMP/PKA pathway activity is reduced, additional nutritional supplementation is provided or if nutrient uptake processes are elevated. Thus, the Serine225 reside plays an important role in the control of Ras2 localisation and activity that allows the cell to co-ordinate nutritional availability with growth and cell division. My thesis highlights that post-translational modifications in regions outside of the highly conserved Ras GTPase domain may be targeted to change cell fate, for example by switching a pro-growth signalling programme to one that drives a growth cessation. This has implications for the development of novel therapeutic approaches for cancers driven by oncogenic Ras proteins.