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J Biol Chem, Vol. 275, Issue 5, 3610-3618, February 4, 2000

The Transcription Factor delta EF1 Is Inversely Expressed with Type II Collagen mRNA and Can Repress Col2a1 Promoter Activity in Transfected Chondrocytes*

Darryl MurrayDagger , Patricia PrechtDagger , Richard BalakirDagger , and Walter E. Horton Jr.§

From the Dagger  Laboratory of Biological Chemistry, Gerontology Research Center, NIA, National Institutes of Health, Baltimore, Maryland 21224 and the § Department of Anatomy, Northeastern Ohio Universities College of Medicine, Rootstown, Ohio 44272

The regulation of Col2a1, which encodes type II collagen, likely results from a balance of both positive and negative proteins. Here we present evidence that the transcription factor delta EF1 participates in the negative regulation of Col2a1 transcription. A deletion analysis suggested that a region between -100 and -307 of the rat Col2a1 gene was required for activity in differentiating chick limb bud mesenchymal cells; however, mutation of a conserved E2 box site in this region actually increased promoter activity. Supershift analysis demonstrated that delta EF1, a known transcriptional repressor, bound to the E2 box in a sequence-dependent manner. Chick limb bud mesenchymal cells, which do not express type II collagen, expressed abundant delta EF1 mRNA, but, following differentiation in micromass culture, delta EF1 mRNA expression was lost. Primary embryonic chick sternal chondrocytes, which express abundant type II collagen, displayed minimal levels of delta EF1 mRNA. The inhibition of Col2a1 transcription following treatment of chick sternal chondrocytes with growth factors was accompanied by increased delta EF1 expression. Overexpression of delta EF1 in differentiated chondrocytes resulted in decreased expression of a reporter construct containing a collagen II promoter/enhancer insert; however, this negative regulation was not dependent on the proximal E2 box. This is the first report of a specific transcription factor involved in the negative regulation of Col2a1.

* 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.

To whom correspondence should be addressed: Dept. of Anatomy, Northeastern Ohio Universities College of Medicine, 4209 State Route 44, Rootstown, OH 44272. Tel.: 330-325-6290; Fax: 330-325-5913; E-mail: wehj@neoucom.edu.

Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
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