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Research Articles: Therapeutics, Targets, and Development

Modulation of the expression of the invasion-suppressor CRMP-1 by cyclooxygenase-2 inhibition via reciprocal regulation of Sp1 and C/EBP

¹ Institute of Biomedical Sciences, Academia Sinica; ² NTU Center of Genomic Medicine, National Taiwan University; ³ Department of Internal Medicine, National Taiwan University Hospital, Taipei, Taiwan; ⁴ School of Cosmeceutics, China Medical University; ⁵ Institutes of Biomedical Sciences and Molecular Biology, National Chung-Hsing University, Taichung, Taiwan

Requests for reprints: Pan-Chyr Yang, Department of Internal Medicine, National Taiwan University Hospital, No. 7, Chung Shan South Road, Taipei, Taiwan. Phone: 886-2-2356-2185; Fax: 886-2-2358-2867. E-mail: pcyang{at}ntu.edu.tw

Abstract

Collapsin response mediator protein-1 (CRMP-1) controls neural development and axonal growth but also acts as a cancer invasion suppressor. In this study, we investigated the transcriptional regulation of CRMP-1 expression. Using a serial deletion strategy, we identified a basal promoter region between nucleotides -100 and -180 in the 5' flanking region of CRMP-1 (nucleotides -1,920 to +50) that contains multiple putative Sp1 and C/EBP sites. Site-directed mutagenesis and deletion analysis revealed that the two C/EBP sites, from nucleotides -122 to -133 and from nucleotides -101 to -113, are the most important regulatory elements. Gel-shift and antibody supershift assays showed that Sp1 protein was also present at this C/EBP site, which overlaps with a Sp1 site. Overexpression of Sp1 decreased CRMP-1 promoter activity and protein expression, whereas overexpression of C/EBP produced the opposite effect. Chromatin immunoprecipitation assays confirmed that Sp1 and C/EBP compete for binding at the overlapping C/EBP and Sp1 sites and reciprocally regulate CRMP-1 expression. Overexpression of cyclooxygenase-2 (COX-2) decreased CRMP-1 mRNA and protein expression. Conversely, the COX-2 inhibitor, celecoxib, induced a dose-dependent increase in CRMP-1 expression. COX-2 inhibition also decreased Sp1-DNA complex formation and inhibited cell invasion. We conclude that transcription of the invasion suppressor, CRMP-1, is reciprocally regulated at the promoter region by C/EBP and Sp1. COX-2 inhibitors increase CRMP-1 expression by inhibiting Sp1-DNA complex formation and enhancing DNA binding of C/EBP at the promoter. [Mol Cancer Ther 2008;7(6):1365–75]

Grant support: National Research Program for Genomic Medicine grants DOH95-TD-G-111-010, DOH95-TG-G-111-012, DOH96-TD-G-111-09, and DOH96-TD-G-111-011.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

Note: C-C. Wu and J-C. Lin contributed equally to this work. T-M. Hong and P-C. Yang codirected the project and contributed equally.

⁶ Supplementary material for this article is available at Molecular Cancer Therapeutics Online (http://mct.aacrjournals.org/).

⁷ http://www.ifti.org/cgi-bin/ifti/Tfsitescan.pl

Received 1/24/08; revised 3/28/08; accepted 3/31/08.