Transposon-mediated, cell type-specific transcription factor recording in the mouse brain

Transcription factors (TFs) play a central role in the regulation of gene expression, controlling everything from cell fate decisions to activity dependent transcription. However, widely-used methods for TF profiling in vivo (e.g. ChIP-seq) yield only an averaged picture of TF binding across all cell types present within the harvested tissue; thus, it is challenging or impossible to determine how the same TF might bind different portions of the genome in different cell types, or even to identify its binding events at all in rare cell types in a complex tissue such as the brain. Here we present a versatile methodology, FLEX calling cards, for the mapping of TF occupancy in specific cell types from heterogenous tissues. In this method, the TF of interest is fused to a hyperactive piggyBac transposase (hypPB), and this bipartite gene is delivered, along with donor transposons, to mouse tissue via a Cre-dependent adeno-associated virus (AAV). The fusion protein is expressed in Cre-expressing cells where it inserts transposon calling cards near to TF binding sites. These transposons permanently mark TF binding events and can be mapped using high-throughput sequencing. Alternatively, unfused hypPB interacts with and records the binding of the super enhancer (SE)-associated bromodomain protein, Brd4. To demonstrate the FLEX calling card method, we first show that donor transposon and transposase constructs can be efficiently delivered to the mouse brain with AAV and that insertion profiles report TF occupancy. Then, using a Cre-dependent hypPB virus, we show utility of this tool in defining cell type-specific TF profiles in multiple cell types of the brain. Finally, we demonstrate utility of FLEX calling cards in longitudinal, integrative recording of the promoter-associated TF, SP1, providing a historical record of SP1 occupancy across time. This approach will enable important cell type-specific studies of TF-mediated gene regulation in the brain and could provide valuable insights into brain development, homeostasis, and disease.