An insulator blocks access to enhancers by an illegitimate promoter, preventing repression by transcriptional interference
Several distinct activities and functions have been described for chromatin insulators, which separate genes along chromosomes into functional units. Here, we describe a novel mechanism of functional separation whereby an insulator prevents gene repression. When thehomieinsulator is deleted from the end of a Drosophilaeven skipped(eve) locus, a flanking P-element promoter is activated in a partialevepattern, causing expression driven by enhancers in the 3’ region to be repressed. The mechanism involves transcriptional read-through from the flanking promoter. This conclusion is based on the following. Read-through driven by a heterologous enhancer is sufficient to repress, even whenhomieis in place. Furthermore, when the flanking promoter is turned around, repression is minimal. Transcriptional read-through that does not produce anti-sense RNA can still repress expression, ruling out RNAi as the mechanism in this case. Thus, transcriptional interference, caused by enhancer capture and read-through when the insulator is removed, repressesevepromoter-driven expression. We also show that enhancer-promoter specificity and processivity of transcription can have decisive effects on the consequences of insulator removal. First, a coreheat shock 70promoter that is not activated well byeveenhancers did not cause read-through sufficient to repress theevepromoter. Second, these transcripts are less processive than those initiated at the P-promoter, measured by how far they extend through theevelocus, and so are less disruptive. These results highlight the importance of considering transcriptional read-through when assessing the effects of insulators on gene expression.