The yeast rRNA gene enhancer does not function by recycling RNA polymerase I and cannot act as a UAS

The mechanism of action of the yeast rRNA gene enhancer was investigated by measuring transcription of an rRNA minigene, cloned into a multicopy plasmid, in transformed yeast. Expression of the minigene was increased when the enhancer was cloned either upstream of or downstream from the minigene. When an enhancer was present both upstream and downstream of the minigene, the upstream element was functionally dominant. The upstream enhancer was active in this construct in the absence of detectable read-through by any RNA polymerase. In a construct containing tandem rRNA minigenes, an enhancer element located between the two promoters activated transcription from both independently. Therefore, the enhancer does not appear to activate transcription by recycling RNA polymerase I molecules to the promoter. The enhancer also failed to activate transcription from the intact promoter of the yeast CYC1 gene, and was unable to functionally substitute for the natural upstream activation sequences (UASs) of this gene. Therefore, the enhancer functions differently to UASs of RNA polymerase II genes, and is probably polymerase-specific.