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

^* To whom correspondence should be addressed. 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]

Key Words: CRMP-1, invasion suppressor, COX-2 inhibitor, Sp1, C/EBP