Gene regulation is fundamental to every aspect of organismal biology. The onset of tissue-specific gene expression in higher organisms depends on the function of genomic elements called distal enhancers. Enhancers activate transcription by binding transcription factors and by communicating their state of occupancy to basal promoters, often located tens to hundreds of kilobases away. How enhancers communicate with their target gene promoters over such genomic distances is one of the greatest mysteries of modern biology.
We propose to address this question by elucidating the dynamic 3D organization of the Drosophila shavenbaby locus in different transcriptional states. The shavenbaby gene encodes a transcription factor that directs the development of cuticular hair-like projections called trichomes. Its embryonic expression is regulated by the combined activities of seven enhancers that are scattered in a 90 Kilobase gene desert upstream of the shavenbaby promoter. We have previously deciphered the regulatory code encoded in three of these enhancers, but it remains unknown how they remotely control shavenbaby expression.
Here we request funds for developing a single-molecule live-imaging system that will allow us to monitor the dynamics of enhancer-promoter communication at the shavenbaby locus and to correlate it to active transcription in live embryos. We will use advanced genetic editing scheme to fluorescently tag the endogenous shavenbaby enhancers, promoter and transcript. Using super-resolution live-imaging we will monitor the relative nuclear localization of these loci in different transcriptional landscapes and states. The proposed experiments will reveal the mechanisms by which distal enhancers communicate to basal promoters to activate tissue-specific gene expression in higher organisms.