Vascular smooth muscle cell-specific regulation of cyclin-dependent kinase inhibitor p21WAF1/Cip1 transcription by Sp1 is mediated via distinct cis-acting positive and negative regulatory elements in the proximal p21WAF1/Cip1 promoter
✍ Scribed by Mary M. Kavurma; Levon M. Khachigian
- Publisher
- John Wiley and Sons
- Year
- 2004
- Tongue
- English
- Weight
- 323 KB
- Volume
- 93
- Category
- Article
- ISSN
- 0730-2312
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✦ Synopsis
Abstract
Smooth muscle cells (SMC) play a central role in common vascular pathologies such as atherosclerosis and restenosis. Understanding the molecular regulation of SMC proliferation at a transcriptional level may provide important clues for the targeted control of vascular hyperplasia. We recently reported the capacity of the transcription factor Sp1 to down‐regulate p21^WAF1/Cip1^ production thereby reducing p21^WAF1/Cip1^–cyclin D1–Cdk4 complex formation and inhibiting vascular SMC proliferation (Kavurma and Khachigian [2003] J. Biol. Chem. 278, 32537–32543). We have now localized the Sp1‐response elements in the p21^WAF1/Cip1^ promoter responsible for p21^WAF1/Cip1^ repression in WKY12‐22 SMCs. The proximal region of the p21^WAF1/Cip1^ promoter contains five distinct Sp1‐binding elements that we have termed A, B, C, D, and E. Electrophoretic mobility shift analysis revealed that SMC nuclear Sp1 interacts with all five Sp1‐binding sites, and each of these sites is critical for Sp1 repression of the p21^WAF1/Cip1^ promoter, since mutation in any one element ablates repression, and in some cases results in activation. In contrast, only elements C, D, and E are bound by Sp1 in endothelial cells. Sp1 overexpression activates the p21^WAF1/Cip1^ promoter in this cell type. Furthermore, mutation in any of these five elements is not sufficient to prevent activation of the p21^WAF1/Cip1^ promoter by Sp1 in endothelial cells. Surprisingly, double mutations of elements C and E facilitates superactivation by Sp1 in both cell types, whereas triple mutations of C, D, and E inactivate the promoter. These findings demonstrate cell type‐specific regulation of p21^WAF1/Cip1^ transcription by Sp1 via distinct cis‐acting positive and negative regulatory elements in the proximal p21^WAF1/Cip1^ promoter. © 2004 Wiley‐Liss, Inc.