Reverse engineering of the transcriptional network downstream of RAF/MAPK signaling

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Mitogen-activated protein kinase (MAPK) signal transduction controls a variety of cellular functions via the induction of hundreds of target genes. While these target programs are known, the transcriptional network that orchestrates the expression of these target genes is not understood. Here we propose to use the technical advances in genome editing, single cell genomics and advances in computational network reconstruction approaches to reverse engineer the transcriptional network downstream of MAPK. Specifically, we will combine different pooled CRISPR approaches with targeted single cell transcriptomes to generate perturbation-response maps for 50 MAPK dependent transcription factors. This data will then be used as input for a network reconstruction approach termed “response logic” that uses the power of answer set solvers to reconstruct networks that agree with the perturbation-response maps. Specifically designed combinatorial perturbations will be used to validate and refine those networks and to identify and validate redundant and synergistic transcription factors.