The Interplay Between Notch Ligands is Critical for Epidermal Sensory Neuronal Specification and Spatial Patterning in the Ascidian Embryo

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During the process of embryonic development, a fertilized egg develops into an organism composed of many different types of cells such as muscle and nerve. A particularly common mechanism that instructs cells what type to become involves communication via proteins on their surface. The Zeller laboratory studies this process in a simple animal model called an ascidian that allows them to observe individual cells and manipulate the functions of their proteins. This project will utilize an approach called optogenetics, in which a specific color of light manipulates the function of these cell surface proteins to alter how cells communicate with one another. By illuminating the ascidian embryo at different times, they can manipulate the process of cell communication precisely to learn how this mechanism operates. This will be one of the first applications of optogenetics in ascidians and they will provide the broader scientific community with the set of tools that they develop. In addition, they will develop learning modules for both undergraduate and graduate-level courses to provide students with an opportunity to use optogenetics. Lastly, they will share this work with the public by developing an activity that demonstrates the principles of optogenetics to children that will be incorporated into our lab’s annual open house presentation at the San Diego State University Marine Laboratory.The ascidian larval peripheral nervous system (PNS) is composed of a series of ciliated sensory neurons that are patterned, in part, by Notch-mediated lateral inhibition. The sensory neurons express two Notch ligands, Delta and Jagged, at different times and in different spatial domains suggesting that more complex Notch signaling, including lateral induction, may play a role in spatial patterning. To investigate this, the Zeller laboratory will utilize an ontogenetically-regulated transcription factor, coupled with tissue-specifically expressed CRISPR reagents to manipulate Notch ligand function with precise spatiotemporal control. By manipulating the function of both Notch ligands individually and simultaneously at different times during PNS development they will tease apart patterning contributions from lateral inhibition and induction. This will be one of the first uses of optogenetics in ascidians and they will provide reagents to the broader scientific community. Using optogenetics-based approaches in other experimental systems will provide a level of precise spatiotemporal regulation that is currently difficult to achieve. They will develop an optogenetics module that will be introduced into one or more advanced undergraduate laboratory courses at San Diego State University and introduce graduate students and postdoctoral fellows to optogenetics by developing a laboratory module in the ascidian section of the Embryology Course at the Marine Biology Laboratory in Woods Hole. Finally, to expose the general public to optogenetics, they will develop an interactive presentation for children that will explain optogenetics principles that will be incorporated into our lab’s annual open house presentation at the SDSU Marine Laboratory.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.