Vascular smooth muscle cell-specific regulation of cyclin-dependent kinase inhibitor p21(WAF1/Cip1) transcription by Sp1 is mediated via distinct cis-acting positive and negative regulatory elements in the proximal p21(WAF1/Cip1) promoter

J Cell Biochem. 2004 Nov 15;93(5):904-16. doi: 10.1002/jcb.20238.


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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Cycle Proteins / genetics
  • Cell Cycle Proteins / metabolism*
  • Cell Line
  • Cyclin-Dependent Kinase Inhibitor p21
  • DNA-Binding Proteins / metabolism
  • Muscle, Smooth, Vascular / cytology*
  • Myocytes, Smooth Muscle / cytology
  • Myocytes, Smooth Muscle / metabolism*
  • Promoter Regions, Genetic / genetics*
  • Protein Binding
  • Protein Kinases / metabolism*
  • Rats
  • Sp1 Transcription Factor / metabolism*
  • Transcription, Genetic*


  • Cdkn1a protein, rat
  • Cell Cycle Proteins
  • Cyclin-Dependent Kinase Inhibitor p21
  • DNA-Binding Proteins
  • Sp1 Transcription Factor
  • Protein Kinases
  • Sp1 kinase