title: A Fully Synthetic Transcriptional Platform for a Multicellular Eukaryote.
abstract: Regions of genomic DNA called enhancers encode binding sites for transcription factor proteins. Binding of activators and repressors increase and reduce transcription, respectively, but it is not understood how combinations of activators and repressors generate precise patterns of transcription during development. Here, we explore this problem using a fully synthetic transcriptional platform in Drosophila consisting of engineered transcription factor gradients and artificial enhancers. We found that binding sites for a transcription factor that makes DNA accessible are required together with binding sites for transcriptional activators to produce a functional enhancer. Only in this context can changes in the number of activator binding sites mediate quantitative control of transcription. Using an engineered transcriptional repressor gradient, we demonstrate that overlapping repressor and activator binding sites provide more robust repression and sharper expression boundaries than non-overlapping sites. This may explain why this common motif is observed in many developmental enhancers.
authors.affiliation: Janelia Research Campus, Howard Hughes Medical Institute, 19700 Helix Drive, Ashburn, VA 20147, USA. Electronic address: crockerj@janelia.hhmi.org.
authors.forename: Justin
authors.initials: J
authors.lastname: Crocker
authors.affiliation: Janelia Research Campus, Howard Hughes Medical Institute, 19700 Helix Drive, Ashburn, VA 20147, USA.
authors.forename: Albert
authors.initials: A
authors.lastname: Tsai
authors.affiliation: Janelia Research Campus, Howard Hughes Medical Institute, 19700 Helix Drive, Ashburn, VA 20147, USA.
authors.forename: David L
authors.initials: DL
authors.lastname: Stern
mesh_terms: D000818:Animals; D001665:Binding Sites; D029721:Drosophila Proteins / Q000235:genetics / Q000378:metabolism; D004331:Drosophila melanogaster / Q000235:genetics*; D004742:Enhancer Elements, Genetic; D009687:Nuclear Proteins; D012097:Repressor Proteins / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic
chemical_list: D029721:Drosophila Proteins; D009687:Nuclear Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; C512295:zld protein, Drosophila
keywords: Drosophila; enhancer; regulatory networks; synthetic biology; transcription