Regulation of the human cyclin-dependent kinase inhibitor p18INK4c by the transcription factors E2F1 and Sp1.
The p18(INK4c) 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. 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 bp 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.
Blais
Alexandre
A
Molecular Endocrinology and Oncology Research Center, Centre Hospitalier de l'Université Laval Research Center, Centre Hospitalier Universitaire de Quebec, Sainte-Foy, G1V 4G2 Quebec, Canada.
Monté
Didier
D
Pouliot
Frédéric
F
Labrie
Claude
C
D000230:Adenocarcinoma; D001483:Base Sequence; D001665:Binding Sites; D001943:Breast Neoplasms; D018797:Cell Cycle Proteins / Q000378:metabolism; D002843:Chromatin / Q000378:metabolism; D050763:Cyclin-Dependent Kinase Inhibitor p18; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors*; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; D050704:E2F4 Transcription Factor; D004791:Enzyme Inhibitors; D005260:Female; D005786:Gene Expression Regulation; D005982:Glutathione Transferase / Q000235:genetics; D006801:Humans; D008969:Molecular Sequence Data; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D017384:Sequence Deletion; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D016385:TATA Box; D014157:Transcription Factors / Q000378:metabolism*; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D025521:Tumor Suppressor Proteins / Q000235:genetics*
C493971:CDKN2C protein, human; D018797:Cell Cycle Proteins; D002843:Chromatin; D050763:Cyclin-Dependent Kinase Inhibitor p18; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D050687:E2F1 Transcription Factor; C493753:E2F1 protein, human; D050704:E2F4 Transcription Factor; C493762:E2F4 protein, human; D004791:Enzyme Inhibitors; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; D005982:Glutathione Transferase; D018844:Cyclin-Dependent Kinases