A high throughput in vivo protein-DNA mapping approach reveals principles of dynamic gene regulation in mammals (RNA-Seq)

Dynamic binding of transcription factors to DNA elements specifies gene expression and cell fate, in both normal physiology and disease. To date, our understanding of mammalian gene regulation has been hampered by the difficulty of directly measuring in vivo binding of large numbers of transcription factors to DNA. Here, we develop a high-throughput indexed Chromatin ImmunoPrecipitation (iChIP) method coupled to massively parallel sequencing to systematically map protein-DNA interactions. We apply iChIP to reconstruct the physical regulatory landscape of a mammalian cell, by building genome-wide binding maps for 29 transcription factors (TFs) and chromatin marks at four time points following stimulation of primary dendritic cells (DCs) with pathogen components. Using over 180,000 TF-DNA interactions in these maps, we derive an initial dynamic physical model of a mammalian cell regulatory network. Our data demonstrates that transcription factors vary substantially in their binding dynamics, genomic localization, number of binding events, and degree of interaction with other factors. Further, many of the TF-DNA interactions at stimulus-activated genes are established during differentiation and maintained in a poised state. Functionally, the TFs are organized in a hierarchy of different types: Cell differentiation factors bind most of the genes and remain largely unchanged during the stimulation. A second set of TFs bind already in the un-stimulated and preferentially target induced genes. A third set consists of TF that bind mainly after the stimuli and target specific gene functions. Together these factors determine the magnitude and timing of stimulus induced gene expression. Our method, which allowed us to map routinely temporal binding profiles of dozens of TFs, provides a foundation for future understanding of the mammalian regulatory code. Overall design: A study of dynamic binding of transcription factors in an immune cell following pathogen stimulation

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Garber M, Yosef N, Goren A, Raychowdhury R, Thielke A, Guttman M, Robinson J, Minie B, Chevrier N, Itzhaki Z, Blecher-Gonen R, Bornstein C, Amann-Zalcenstein D, Weiner A, Friedrich D, Meldrim J, Ram O, Cheng C, Gnirke A, Fisher S, Friedman N, Wong B, Bernstein BE, Nusbaum C, Hacohen N, Regev A, Amit I