Sensing and processing information through signaling cascades is an essential part of cellular life. A few signaling cascades such as the MAP kinase and Hippo pathways are ubiquitous among eukaryotes yet perform different functions across organisms. Although these pathways are well-studied, how they evolve to take on new functions and adapt to new inputs remains poorly understood. The Mitotic Exit Network (MEN), a Ras-like GTPase signaling cascade and yeast homolog of the Hippo pathway, provides a unique opportunity to study this question. In the MEN, the same core signaling components operate in distinct manners under different developmental trajectories. During yeast mitosis which occurs through an asymmetric cell division called budding, the MEN is scaffolded onto the spindle pole bodies (SPB, the yeast equivalent of centrosomes) and responds to spindle position through its GTPase Tem1. During meiosis, where budding is suppressed and thus no need to sense spindle position, MEN signaling is no longer organized at the SPBs, and it is unclear whether Tem1 is still required for MEN activation and what signal it may respond to. To understand the adaptation of the MEN under distinct cellular contexts, this proposal will test the hypothesis that this adaptation is enabled partially by different activation mechanisms of the MEN kinase Cdc15, the effector kinase of Tem1, between mitosis and meiosis (Aim 1). In contrast to the drastic change in spatial organization of the MEN core components between mitosis and meiosis, the effector protein of the MEN, the phosphatase Cdc14, remains sequestered in the nucleolus prior to activation both in mitosis and meiosis. In fact, this nucleolar localization of Cdc14 is conserved from yeast to human. Sequestration of Cdc14 in the nucleolus could function either 1) to ensure tight inhibition of Cdc14?s phosphatase activity prior to activation or 2) to localize Cdc14 to dephosphorylate specific substrates in the nucleolus. To uncover the selection pressure that maintains this conserved nucleolar localization of Cdc14, this proposal will examine these two hypotheses by sequestering Cdc14 elsewhere in the cell and characterize the consequences first in yeast and then in mammalian cells (Aim 2). The experiments within both aims will be initiated during the K99 phase which also includes training of the candidate on new experimental systems such as yeast meiosis and mammalian cells, as well as the development and implementation of quantitative microscopy, proximity labeling and optogenetics. Furthermore, the candidate has assembled an outstanding mentor team to both advise her scientifically to facilitate progress of the project and prepare her for the transition to an independent investigator. Together, this proposal will create a strong foundation for an independent research career in understanding the evolution/adaptation and spatial organization of cellular signaling.

Public Health Relevance

How signaling pathways evolve to adapt to new inputs and why certain spatial organization of signaling components are conserved across species while others are rewired remain poorly understood. This proposal will address these questions using the yeast Mitotic Exit Network (MEN) as a model system taking advantage of the intriguing observation that the core components of the MEN are rewired from mitosis to meiosis to adapt to different developmental inputs while the nucleolar localization of the MEN effector, the protein phosphatase Cdc14, is evolutionarily conserved from yeast to human. Results from this work will advance our understanding of the evolution and spatial organization of cellular signaling, whose dysfunction underlies a number of diseases such as cancer.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Career Transition Award (K99)
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Special Emphasis Panel (ZGM1)
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Gaillard, Shawn R
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Massachusetts Institute of Technology
Veterinary Sciences
United States
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