The role of Ras modulators and feedback control mechanisms on the establishment of oscillations in the Ras/cAMP/PKA pathway in S. cerevisiae

In S. cerevisiae, the Ras/cAMP/PKA pathway is involved in the regulation of metabolism and cell cycle progression. The synthesis of cAMP is mediated by adenylate cyclases, whose activity is controlled by Ras protein, a GTPase that cycles between an active GTP-bound state and an inactive GDP-bound state. Ras is positively regulated by Cdc25 (a Ras-GEF) and negatively regulated by Ira (a Ras-GAP). cAMP activates PKA, which in turn exerts several feedback controls in the pathway, while the two phosphodiesterases Pde1 and Pde2 control the signal termination through the degradation of cAMP. Experimental and computational investigations of this pathway are complicated by the interplay between all these regulation mechanisms. To understand the role of feedback controls, we defined a mechanistic model of the Ras/cAMP/PKA pathway and studied the dynamics of its pivotal components. Simulations and parameter sweep analysis of this model indicate that stable oscillatory regimes of cAMP can be established when the negative feedback on Ira proteins is activated, and that this dynamics is regulated by the balance between the activities of Cdc25 and Ira. Our analysis highlights that also the guanine nucleotide pools, which concurrently regulate the switch cycle of Ras, are able to control the transition between steady states and oscillations of cAMP when the ratio GTP/GDP drops to a critical level. In addition, we studied the influence of phosphodiesterases, suggesting that the feedback on Pde1 can regulate the oscillatory regime of cAMP independently from the presence of Pde2. This multi-level regulation of the pathway might function to extend the regulatory span of the system in the cell, and might act as a tuning mechanism of downstream targets, in line to the recently suggested hypothesis of “frequency-modulated” gene regulation. Our analysis shows, in a quantitative way, how the coupling between different feedback mechanisms and the molecular levels of Ras modulators (Cdc25/Ira2, GTP/GDP) influence the form and the frequency of oscillations. The model allows to make predictions on the conditions that lead to the insurgence of oscillations in the pathway, and represents a useful tool to better understand the experimental data or to plan focused experiments. Further work is in progress to analyse the additional effects of the Gpr1/Gpa2 pathway, that works in parallel with Ras to activate PKA in response to various sugar ligands.