Engineering tyrosine kinase signaling in yeast

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Ricardo Almeida, Andrew Horwitz, Joan Garbarino, Jeff Johnson, Erwin Schoff, Rune Linding, Nevan Krogan, Wendell Lim

University of California San Francisco, United States

Complex cellular behaviors are orchestrated by networks of signal transduction pathways that process sensory information to execute specific physiological functions. Tyrosine kinase (TyrK) signaling is an example of a recently evolved signaling system that has become an essential part of modern metazoan cell biology. Phosphotyrosine (pTyr) signaling circuits mediate cell-to-cell communication responsible for controlling cell migration, differentiation, hormone response and immune defense. Its underlying architecture is a combination of three components: the writer (Tyrosine kinase, TyrK), the reader (SH2 domain, specifically binds phosphotyrosine peptides) and the eraser (protein tyrosine phosphatase, PTP). This modular, versatile design has enabled cells to develop vast set of complex signaling networks and provides us with a rich toolkit with which to engineer novel signaling circuits to artificially control cellular behavior.

In order to study the design principles of pTyr signaling circuits in a naive cellular environment, we are introducing pTyr signaling to the budding yeast Saccharomyces cerevisiae, which lacks endogenous TyrK or SH2 domain containing proteins. We are developing artificial phosphorylation cascades using designed TyrK and SH2 substrates towards building novel signal transduction circuits with interesting properties. Through a combination of proteomics, cell biology and in vitro biochemistry we have uncovered the mechanistic basis for growth defects resulting from Src TyrK expression in yeast. Our results highlight the engineering challenges of developing an orthogonal signaling system in a naive host as well as provide insights into the emergence of a novel signaling currency in eukaryotic evolution.