An interplay of Sp1, GKLF and CREB-2 controls human Pre-alpha-Inhibitor gene (ITIH3) transcription.
Pre-alpha-Inhibitor is a plasma protease inhibitor and a heterodimeric molecule whose one polypeptide chain is encoded by the ITIH3 gene. In order to understand the expression of this protein that is regulated in health and disease, we have analyzed the 5' flanking region of ITIH3, specifically focussing on its proximal promoter. A combination of methods including wild-type (wt) or mutant promoter linked to a reporter cat gene, co-transfections of cat constructs with expression plasmids for nuclear factors and electrophoretic mobility shift assays revealed that two antagonistic sets of regulatory elements and nuclear proteins are critical for the activity of this promoter. Indeed, several overlapping Sp1/Sp3-binding sites are required for a sustained activity. However, a tripartite complex including CREB-2 and two molecules of the gut-enriched, Krüppel-like factor cooperate to bind to an upstream area whose 3' end overlaps the Sp1-binding sites. The resulting competition between this tripartite complex and Sp1 results in impaired occupancy of Sp1-binding sites by Sp1 and a consequent reduction in ITIH3 transcription. Competition between Sp1 and a Krüppel-like factor for GC-rich sites has been previously reported, but this is the first description of an elaborate tripartite cooperation of two Krüppel-like factors and CREB as a key step in such a competition.
Ruminy
Philippe
P
INSERM Unit 519, Faculté de Médecine-Pharmacie, Institut Fédératif de Recherches Multidisciplinaires sur les Peptides, 22 Bvd Gambetta, 76183 Rouen cedex, France.
Rouet
Philippe
P
Salier
Jean-Philippe
JP
D051701:Activating Transcription Factor 4; D001665:Binding Sites / Q000235:genetics; D045744:Cell Line, Tumor; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D005786:Gene Expression Regulation; D006801:Humans; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; D009841:Oligonucleotides / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D011498:Protein Precursors / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D016329:Sp1 Transcription Factor / Q000235:genetics / Q000378:metabolism; D051705:Sp3 Transcription Factor; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic; D014162:Transfection; D014361:Trypsin Inhibitors / Q000235:genetics*
C497109:ATF4 protein, human; D004268:DNA-Binding Proteins; C000715467:KLF4 protein, human; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; D009841:Oligonucleotides; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; C497121:SP3 protein, human; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; D014361:Trypsin Inhibitors; C061151:pre-alpha-trypsin inhibitor; D051701:Activating Transcription Factor 4; D051705:Sp3 Transcription Factor; D015500:Chloramphenicol O-Acetyltransferase