ANALYSIS OF SCAFFOLD-PROTEIN MEMBRANE RECRUITMENT DYNAMICS DURING PHEROMONE RESPONSE IN Saccharomycfes cerevisiae

VICTORIA REPETTO; ALAN BUSH; ALEJANDRO COLMAN LERNER

Potrero de Funes

Congreso; SAIB 47th Annual Meeting Argentine Society of Biochemistry and Molecular Biology XLVII Reunión Anual Argentina de Investigación en Bioquímica y Biología Molecular; 2011

Society of Biochemistry and Molecular Biology XLVII Reunión Anual Argentina de Investigación en Bioquímica y Biología Molecular

The yeast mating pheromone response pathway is a prototypical example of a signaling pathway that uses a scaffold protein. The scaffold, Ste5, has several interaction domains, some of which mediate essential steps necessary for the activation of its associated three-tiered MAPK cascade, while others function in higher-order regulatory behaviors, such as feedback phosphorylation by the MAPK Fus31 and inhibitory phosphorylation by the cell-cycle dependent kinase2. This work is aimed at determining the role of Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. dependent kinase2. This work is aimed at determining the role of Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. dependent kinase2. This work is aimed at determining the role of Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. dependent kinase2. This work is aimed at determining the role of Ste5 interaction domains in the dynamics of the pheromone response. To do this, we followed in real-time recruitment of Ste5 to the membrane in single cells upon pheromone treatment, analyzing the redistribution of a Ste5 YFP fusion with a confocal microscope. We obtained quantitative measurements from single-cells using Cell- ID coupled data analysis with R, following a method previously developed in our lab3. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular domains in pathway dynamics. Mutations at Ste5 regulatory sites, mimicking constitutive (de)phosphorylation, produced changes in the dynamic of membrane binding, suggesting the importance of such modifications for the finetuning of the response. We measured several Ste5 mutants to assess the role of Ste5 modular