Molecular recording to predict cell fate decisions and animal behavior

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Abstract The brain is remarkably complex, and our understanding of this organ is still in its infancy. Many fundamental questions about brain development and function remain. Single cell genomics promises new ways to answer these questions, but nearly all single-cell methods share shortcoming – cells are destroyed when their molecular states are measured. This “destruction upon observation” makes it impossible to correlate molecular events that have occurred in the past with the final outcomes: if you destroy an intermediate progenitor in the brain to profile its epigenome, you have no way to know what cell fate the progenitor would have adopted. Likewise, profiling an animal after an experience is going to conflate the molecular consequences of the experience with any molecular predispositions that were there before exposure. And if you profile before the experience, you have no way of know of the animal's outcome – e.g. if the animal would have been a learner or not. This shortcoming is inherent to nearly all existing genomic methods and impedes a wide variety of interesting analyses. We have recently developed a platform technology, single-cell `Calling Cards' (CC) that uses transposons to capture molecular events at one instant in time and then read them out at a later time with single cell resolution. We now propose to build upon this foundation to develop a robust, easy to use technology platform to record molecular events at single cell resolution in the mouse brain and connect them with cell fate decision, behavioral outcomes, neuronal activity profiles, and anatomical location. We will develop a set of widely applicable reagents and methods, apply them to a series of high-impact “test cases” to demonstrate their utility, and rapidly distribute them to the broader community.