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Volume 271, Number 43, Issue of October 25, 1996 pp. 26717-26723
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.

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Transcriptional Regulation of the Human 2(I) Collagen Gene
COMBINED ACTION OF UPSTREAM STIMULATORY AND INHIBITORY CIS-ACTING ELEMENTS

(Received for publication, April 30, 1996, and in revised form, June 25, 1996)

From the Department of Medicine, Division of Rheumatology, Medical University of South Carolina, Charleston, South Carolina 29425-2229

This study identifies three regions of the human 2(I) collagen promoter involved in the binding of nuclear factors. These regions include sequences from 173 to 155 (footprint I), 133 to 119 (footprint II), and 101 to 72 (footprint III). A novel positive cis-element containing a TCCTCC motif was identified within footprint II. In addition, we demonstrated that a pyrimidine-rich region within footprint I is a binding site for a transcriptional repressor, and a CCAAT motif within footprint III is a binding site for a transcriptional activator. Comparative functional analysis of the cis-acting elements within the proximal 350 base pairs of this promoter, including previously characterized Sp1 binding sites at 300, indicates that constitutive activity of this promoter is regulated equivalently by the three positive cis-acting elements at 300, 125, and 80. Mutations in the repressor site at 160 increase constitutive activity by 4-6-fold. However, simultaneous mutations of the repressor site and the cis-regulatory element at either the 300 or 125 sites result in no increase in constitutive transcription activity suggesting interaction between the activators and repressor elements. In contrast, simultaneous mutation of the CCAAT motif and the repressor site results in about a 4-fold increase, suggesting that activation via the CCAAT motif may be independent of this repressor.

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