Reconstitution of enhancer function in paternal pronuclei of one-cell mouse embryos

How chromatin-mediated transcription regulates the beginning of mammalian d evelopment is currently unknown. Factors responsible for promoter repressio n and enhancer-mediated relief of this repression are not present in the pa ternal pronuclei of one-cell mouse embryos but are present in the zygotic n uclei of two-cell embryos. Here we show that coinjection of purified histon es and a plasmid-encoded reporter gene into the paternal pronuclei of one-c ell embryos at a specific histone-DNA concentration could recreate the beha vior observed in two-cell embryos: acquisition of promoter repression and s ubsequent relief of this repression either by functional enhancers or by hi stone deacetylase inhibitors. Furthermore, the extent of enhancer-mediated stimulation in one-cell embryos depended on the acetylation status of the i njected histones, on the treatment of embryos with a histone deacetyllase i nhibitor, and on the developmentally regulated appearance of enhancer-speci fic coactivator activity. The coinjected plasmids in one-cell embryos also exhibited chromatin assembly, as determined by a supercoiling assay. Thus, injection of histones into one-cell embryos faithfully reproduced the chrom atin-mediated transcription observed in two-cell embryos. These results sug gest that the need for enhancers to stimulate promoters through relief of c hromatin-mediated repression occurs once the parental genomes are organized into chromatin. Furthermore, we present a model mammalian system in which the role of individual histones, and particular domains within the histones that are targeted in enhancer function, can be examined using purified mut ant histones.