A germline- and promoter-independent strategy to gain access to all cell types in the brain

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Project summary The heterogeneity from the vast number of cell types in the brain presents a major challenge in our understanding of how brain works and in our treatment of neurological disorders. With the amazing advances in high throughput sequencing technology, our knowledge on the molecular makeup of the myriad cell types in the brain has reached an unprecedented level. However, tools that allow us to easily study the functions of any cell types of our choice are lagging. The goal of our proposed research is to develop technology to generate such tools. In order to target a specific cell type, current approaches typically depend on genetically modified animal models, which is laborious, costly, and low throughput, or well-defined and small cell-type-specific promoters, which still remain to be difficult to isolate. Here we propose to develop an AAV-based Cell-Type-Specific Targeting (ACTSTar) system to enable easy access to any cell type in the brain in any species that is independent of germline modification and promoter isolation and characterization. We plan to generate a library of ACTSTar drivers to target 50 different brain cell types in the mouse and marmoset. In addition, to reduce off-target integration and neuroinflammation associated with long-term Cas9 expression, especially in long-term experiments in primates, we will develop a novel AAV-capsid mediated Cas9 mRNA delivery system for transient Cas9 expression. This project will generate the much-needed tools for easy access to any brain cell types in multiple species. Our research goal will be facilitated by the complementary expertise of the MPI team in genome editing and developing safe Cas9 delivery methods (Lu), neural circuit functions of the mouse brain (Lin), and visual functions using non-human primates (Hu).