Co-occupancy identifies transcription factor co-operation for axon growth.

Transcription factors (TFs) act as powerful levers to regulate neural physiology and can be targeted to improve cellular responses to injury or disease. Because TFs often depend on cooperative activity, a major challenge is to identify and deploy optimal sets. Here we developed a bioinformatics pipeline, centered on TF co-occupancy of regulatory DNA, and used it to predict factors that potentiate the effects of pro-regenerative Klf6 in vitro. High content screens of neurite outgrowth identified cooperative activity by 12 candidates, and systematic testing in a mouse model of corticospinal tract (CST) damage substantiated three novel instances of pairwise cooperation. Combined Klf6 and Nr5a2 drove the strongest growth, and transcriptional profiling of CST neurons identified Klf6/Nr5a2-responsive gene networks involved in macromolecule biosynthesis and DNA repair. These data identify TF combinations that promote enhanced CST growth, clarify the transcriptional correlates, and provide a bioinformatics approach to detect TF cooperation.