Molecular and Cellular Biochemistry

, Volume 116, Issue 2, pp 119–129

An AP-1-like motif in the first intron of human Proα1(I) collagen gene is a critical determinant of its transcriptional activity

  • Hitoshi Katai
  • Jane D. Stephenson
  • Carl P. Simkevich
  • James P. Thompson
  • Rajendra Raghow


The first intron of the human Proα1(I) collagen gene contains an orientation-dependent enhancer composed of both positive and negative cis-acting elements involved in the transcriptional regulation of this gene. Deletion of a 360 bp Sau 3A intronic fragment spanning nucleotide + 494 to + 854 (S360) resulted in dramatic down-regulation of pCOL-KT (Thompson et al., J Biol Chem 266: 2549–2556, 1991). Using a DNaseI protection assay, we demonstrate a single footprint located at + 590 to + 615 in the S360 fragment; nuclear extracts prepared from mesenchymal and nonmesenchymal cells exhibited similar binding characteristics. A double stranded oligonucleotide representing a consensus Ap-1 binding sequence competed with S360 for binding. In contrast to what occurred in response to S360 deletion which was always accompanied by reduced expression, the deletion of the Ap-1 binding site (+ 598 to + 604) caused either increased or decreased expression of the reporter gene depending on the target cell. Site-directed mutations in the Ap-1-like cis-element of Proα1(I) were also tested in transient expression assays. Consistent with the paradoxical results of Ap-1 deletion, we observed that the functional consequences of mutations in the Ap-1 site also varied in different cells. In A204 cells, one point mutation, which resulted in the loss of protein binding to S360, led to increased CAT activity while another point mutant, which retained binding of the Ap-1 like trans-acting factor(s), showed decreased CAT expression. The effects of these two mutations in the HFL-1 cells were exactly opposite of what was seen for A204 cells. Based on these observations, we postulate that the Ap-1 site plays a critical role in the transcriptional activity of the human Proα1(I) gene. The implications of an apparently dual mode of regulation through a single cis-regulatory element are discussed. (Mol Cell Biochem 118: 119–129, 1992)

Key words

collagen transcription intronic Ap-1 fos jun trans-acting factors 


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Copyright information

© Kluwer Academic Publishers 1992

Authors and Affiliations

  • Hitoshi Katai
    • 1
    • 2
  • Jane D. Stephenson
    • 1
    • 2
  • Carl P. Simkevich
    • 1
    • 2
  • James P. Thompson
    • 1
    • 2
  • Rajendra Raghow
    • 1
    • 2
  1. 1.Departments of Medicine and PharmacologyUniversity of TennesseeMemphis
  2. 2.Research Service (151)Veterans Affairs Medical CenterMemphisUSA

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