COMPETITION ANALYSIS OF THE HUMAN α2(I) COLLAGEN PROMOTER USING SYNTHETIC OLIGONUCLEOTIDES

概要

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Previous studies have identified four cis-acting elements which mediate the basal transcriptional activity of the human α2(I) collagen gene. One of these elements, a pyrimidine-rich region (TCCCCC motif), was shown to be a repressor site, and the other three elements were shown to be activator sites. Furthermore, the repressor site and two of the activator sites were found to constitute binding sites for the transcription factors Sp1 and Sp3. In this study, we further detrmined the affinity and specificity of the binding of Sp1 and Sp3 to the human α2(I) collagen promoter and investigated the function of the pyrimidine-rich region which contains the TCCCCC motif. Competitive analyses using DNA mobility shift assays showed that the TCCCCC motif which constitutes the repressor site abolished the binding of Sp1 or Sp3 to the GC-boxes or TCCTCC motif, but not the binding of CCAAT-binding factor. The affinity of Sp1 or Sp3 for the TCCTCC motif was shown to be greater than that of Sp1 consensus oligonucleotides. In vitro transcription analysis revealed that the addition of each activator site oligonucleotide or repressor oligonucleotide had an inhibitory effect on the transcription of the collagen gene. These results suggest that the repressor site regulates the transcription of the collagen gene by taking away Sp1 or Sp3 from the activator sites.
日本結合組織学会の論文