Ets regulation of the erbB2 promoter

Evaluating the chromatinized erbB2 gene in nuclei from breast cancer cells expressing varying levels of ErbB2 transcripts, we identified a nuclease-se nsitive site within a 0.22 kb region of maximum enhancer activity centered over a conserved 28 bp polypurine(GGA)-polypyrimidine(TCC) mirror-repeat an d an adjacent essential Ets binding site (EBS), Promoter footprinting with nuclear extracts reveals an intense Ets hypersensitivity site at the EBS wh ose degree of intensity correlates with the Level of cellular ErbB2 express ion. In vitro mapping assays show that the supercoiled erbB2 promoter forms an internal tripler structure (Hr-DNA) at the mirror-repeat element. Mutat ions preventing Hr-DNA formation can enhance erbB2 promoter activity in hum an breast cancer cells, a result consistent with previous demonstration tha t Ets-erbB2 promoter complexes cannot form when the mirror-repeat is engage d in tripler binding, and new results suggesting that Ets binding induces s evere promoter bending that may restrict local tripler formation. In additi on to previously described erbB2-regulating breast cancer Ets factors (PEA3 , ESX/Elf-3), Elf-1 is now shown to be another endogenously expressed Ets c andidate capable of binding to and upregulating the erbB2 promoter. Given c urrent strategies to transcriptionally inhibit ErbB2 overexpression, includ ing development of novel erbB2 promoter-targeted therapeutics, an EBS-targe ted approach is presented using chimeric Ets proteins that strongly repress erbB2 promoter activity.