Investigating FGF Signaling Dynamics in migrating cells

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Project Summary Signaling dynamics are tightly regulated in space and time to control cell migration, proliferation, and differentiation. A comprehensive understanding of how extracellular inputs influence downstream signaling and cell behaviors will first require the ability to visualize key endogenous signaling proteins, cell architectures, and cytoskeletal regulators in vivo. Second, it will be essential to utilize precise tools to manipulate protein localization and signaling activity to reveal mechanisms. To explore how secreted signaling proteins disperse and modulate cell behavior, we are using a type of C. elegans muscle progenitors (SMs) as a tractable model for in vivo cell biology. The SMs migrate from near the tail of the worm to the center during larval development and are a classical system to investigate cell migration mechanisms. SM migration requires Fibroblast Growth Factor (FGF) signaling, which has been hypothesized to act as a long-range chemoattractant. However, the key molecular and cell biological mechanisms that translate FGF signaling into directed migration are not known. FGFs are often thought of as diffusible signaling proteins that can form gradients through free, extracellular dispersal, but this phenomenon has never been demonstrated for an endogenous FGF. To investigate how FGF proteins move between cells to regulate SM migration, I have tagged the endogenous FGF ligand and receptor involved in SM migration with fluorescent proteins and imaged their localization in vivo. Unexpectedly, I did not observe a gradient of FGF protein during SM migration, but instead observed low levels of FGF expression in a line of ventral midline cells that are located near migrating SMs. The proposed work builds on these findings and other new tools that I have made to visualize and manipulate key FGF pathway proteins in order to understand how FGF signaling regulates cell migration. Aim 1 will characterize endogenous FGF dynamics in a living animal and test how FGF moves between cells in vivo using genome engineering and live imaging approaches. Aim 2 will investigate how migrating cells respond to extracellular FGF by testing the extent to which FGF is a permissive or instructive signal, characterizing spatial organization of intracellular signaling downstream of FGF, and testing functions for localized pathway activation using optogenetics. These experiments will provide novel insights into how secreted signaling proteins move between cells and how migrating cells interpret and respond to dynamic extracellular cues.