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Targeting the Ets Binding Site of the HER2/neu Promoter with Pyrrole-Imidazole Polyamides

Chiang, Shu-Yuan and Bürli, Roland W. and Benz, Chris C. and Gawron, Loretta and Scott, Gary K. and Dervan, Peter B. and Beerman, Terry A. (2000) Targeting the Ets Binding Site of the HER2/neu Promoter with Pyrrole-Imidazole Polyamides. Journal of Biological Chemistry, 275 (32). pp. 24246-24254. ISSN 0021-9258. https://resolver.caltech.edu/CaltechAUTHORS:CHIjbc00

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Abstract

Three DNA binding polyamides (1-3) were synthesized that bind with high affinity (Ka = 8.7·10^9 M^-1 to 1.4·10^10 M^-1) to two 7-base pair sequences overlapping the Ets DNA binding site (EBS; GAGGAA) within the regulatory region of the HER2/neu proximal promoter. As measured by electrophoretic mobility shift assay, polyamides binding to flanking elements upstream (1) or downstream (2 and 3) of the EBS were one to two orders of magnitude more effective than the natural product distamycin at inhibiting formation of complexes between the purified EBS protein, epithelial restricted with serine box (ESX), and the HER2/neu promoter probe. One polyamide, 2, completely blocked Ets-DNA complex formation at 10 nM ligand concentration, whereas formation of activator protein-2-DNA complexes was unaffected at the activator protein-2 binding site immediately upstream of the HER2/neu EBS, even at 100 nM ligand concentration. At equilibrium, polyamide 1 was equally effective at inhibiting Ets/DNA binding when added before or after in vitro formation of protein-promoter complexes, demonstrating its utility to disrupt endogenous Ets-mediated HER2/neu preinitiation complexes. Polyamide 2, the most potent inhibitor of Ets-DNA complex formation by electrophoretic mobility shift assay, was also the most effective inhibitor of HER2/neu promoter-driven transcription measured in a cell-free system using nuclear extract from an ESX- and HER2/neu-overexpressing human breast cancer cell line, SKBR-3.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1074/jbc.M000820200DOIArticle
ORCID:
AuthorORCID
Dervan, Peter B.0000-0001-8852-7306
Additional Information:© 2000 The American Society for Biochemistry and Molecular Biology, Inc. Received for publication, January 31, 2000, and in revised form, May 8, 2000. Originally published In Press as doi:10.1074/jbc.M000820200 on May 18, 2000. This study was supported in part by NCI, National Institutes of Health Grant CA16056 (to T.A.B.), National Institutes of Health Grant GM51747 (to P.B.D.) American Cancer Society Grant DHP 158 (to T.A.B.), and United States Army Medical Research Grants BC960313 (to S.Y.C.), CA36773, and CA44768 (to C.C.B.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. [] Recipient of Swiss National Foundation and the "Novartis Stiftung, vormals Ciba-Geigy-Jubiläums-Stiftung" fellowships.
Funders:
Funding AgencyGrant Number
NIHCA-16056
NIHGM-51747
American Cancer SocietyDHP 158
U.S. ArmyBC960313
U.S. ArmyCA36773
U.S. ArmyCA44768
Issue or Number:32
Record Number:CaltechAUTHORS:CHIjbc00
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:CHIjbc00
Official Citation:Shu-Yuan Chiang, Roland W. Bürli, Chris C. Benz, Loretta Gawron, Gary K. Scott, Peter B. Dervan, and Terry A. Beerman Targeting the Ets Binding Site of the HER2/neuPromoter with Pyrrole-Imidazole Polyamides J. Biol. Chem. 2000 275: 24246-. doi:10.1074/jbc.M000820200
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:10741
Collection:CaltechAUTHORS
Deposited By: Archive Administrator
Deposited On:05 Jun 2008
Last Modified:12 Dec 2019 17:07

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