Cell-free genomics: transcription factor interactions in reconstituted naïve embryonic chromatin

Abstract

Extracts from Drosophila preblastoderm embryos (DREX) form the basis of a powerful in vitro chromatin reconstitution system that assembles entire genomes into complex chromatin with physiological nucleosome spacing and polymer condensation. As the zygotic genome has not yet been activated in preblastoderm embryos, the reconstitution extract lacks endogenous transcription factors (TFs) and the RNA polymerase machinery. At the same time, it contains high levels of ATP-dependent nucleosome sliding enzymes that render the reconstituted chromatin dynamic. The naïve chromatin can be used to determine the intrinsic DNA binding properties of exogenous, usually recombinant TFs (or DNA binding proteins in general) in a complex chromatin context. Recent applications of the system include the description of cooperation and competition of Drosophila pioneer TFs for composite binding sites, and the characterization of nucleosome interactions of mammalian pioneer TFs in the heterologous system.

Keywords: Drosophila preblastoderm embryo; chromatin immunoprecipitation; nucleosome binding; nucleosome remodelling; pioneer transcription factor.