EAGER: Functional Integration of Synthetic Kinases into Endogenous Signaling Networks

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Cells can acquire new genes by natural mechanisms (e.g. horizontal gene transfer) or by artificial means (e.g. genetic engineering). We are beginning to understand how newly-acquired genes are integrated into the host's genome, but the evolutionary mechanisms by which the proteins produced by these genes become functionally incorporated into the cell's regulatory networks remain largely unknown. In this project, synthetic biology and directed evolution approaches will be used to recreate and analyze in the laboratory this network rewiring process. Using yeast that have been engineered with signaling modules taken from the mammalian mitogen-activated protein kinase cascade, evolutionary strategies will be applied in the laboratory to reward individual yeast cells that acquire mutations resulting in functional utilization of these introduced mammalian proteins. During this selection process, genome sequencing will be performed to characterize the mutations that occur and to reconstruct the evolutionary processes that result in functional rewiring of the host's signaling networks. Broader Impacts: The scientific principles underlying this research will be used to train graduate and undergraduate students, including those from underrepresented minorities. Modules on evolution in synthetic biology will be added to an undergraduate course taught by the principal investigator. Undergraduates will also have the opportunity to learn about synthetic biology through the International Genetically Engineered Machine (iGEM) competition. Additional outreach will be targeted to high school teachers and high school students through summer training programs. The insights gained from this project will have relevance for understanding the emergence of antibiotic resistance in microbes, the cultivation and properties of genetically modified crops, and the design of novel synthetic biology devices.