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Papers In Press, published online ahead of print June 20, 2002
J. Biol. Chem, 10.1074/jbc.M204554200
Submitted on May 9, 2002
Revised on June 19, 2002
Accepted on June 19, 2002
Molecular Endocrinology and Oncology Research Center, CHUL Research Center, Sainte-Foy, Qc G1V 4G2
Corresponding Author: Claude.Labrie{at}crchul.ulaval.ca
The p18INK4c cyclin-dependent kinase inhibitor is an important regulator of cell cycle progression and cellular differentiation. We and others found that overexpressed E2F proteins up-regulate p18 expression. In order to better understand this phenomenon we performed a functional analysis of the human p18 promoter. Deletion studies revealed that the E2F-responsive elements of the promoter are located within 131 base pairs upstream of the transcription start site. This region contains putative Sp1 and E2F binding sites. Mutational inactivation of these elements revealed that the Sp1 sites were important for the basal activity of the promoter, but could also mediate the effects of E2F1 on the p18 promoter. Moreover, we found that E2F1 and Sp1 can synergistically enhance the activity of the proximal p18 promoter. Gel shift analyses using p18 promoter derived probes led to the identification of several multiprotein complexes that were found to contain different combinations of E2F proteins and/or Sp1. Recombinant E2F1 was also capable of binding to the E2F binding sites. Chromatin immunoprecipitation experiments demonstrated that E2F1 and E2F4 associate with the p18 promoter in unperturbed cells. Based on these findings, we conclude that E2F proteins and Sp1 play an important role in the control of p18 expression.
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