Control of the tissue inhibitor of metalloproteinases-1 promoter in culture-activated rat hepatic stellate cells: regulation by activator protein-1 DNA binding proteins

Hepatology. 1999 Mar;29(3):839-48. doi: 10.1002/hep.510290333.


In the injured liver hepatic stellate cells (HSCs) undergo a dramatic phenotypic transformation known as "activation" in which they become myofibroblast-like and express high levels of the tissue inhibitor of metalloproteinase 1 (TIMP-1). HSC activation is accompanied by transactivation of the TIMP-1 promoter. Truncation mutagenesis studies delineated a minimal active promoter consisting of nucleotides -102 to +60 relative to the major start site for transcription. Removal of an AP-1 site located at nucleotides -93 to -87 caused almost a complete loss of promoter activity. Analysis of AP-1 DNA binding activities during culture activation of HSCs initially indicated transient expression of proteins capable of forming a low mobility AP-1 DNA binding complex (LMAP-1). LMAP-1 was maximally induced at 24 hours of culture and then fell to undetectable levels at 120 hours. Western blot studies showed that both c-Fos and c-Jun underwent similar transient inductions. These temporal changes in c-Fos and c-Jun activities were unexpected because TIMP-1 mRNA expression is not detected in HSCs until culture day 3 to 5 and is thereafter sustained at a high level. Previous work in other cell lineages has established a key role for Pea3 binding proteins (Ets-1) in AP-1 mediated transactivation of the TIMP-1 promoter. We show that HSCs express relatively low levels Ets-1 and Ets-2 and show that mutagenesis of the Pea3 DNA binding site in the TIMP-1 promoter has less than a twofold effect on its activity in activated HSCs. Further analysis of AP-1 DNA binding activities in 7- to 14-day culture activated HSCs led to the discovery of high mobility AP-1 complexes (HMAP-1). HMAP-1 DNA binding activities were sequence specific with respect to AP-1 and absent from freshly isolated HSCs. Supershift EMSA and Western blot studies identified JunD, Fra2, and FosB as potential components of the HMAP-1. Mutations of the AP-1 site of the TIMP-1 promoter that prevented formation of HMAP-1 caused a 70% loss of activity in transfected activated HSCs. Taken together the data indicate that sustained upregulation of TIMP-1 gene expression may be at least partially controlled by a novel AP-1 dependent regulation of TIMP-1 promoter activity.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Base Sequence
  • Binding Sites / physiology
  • Cells, Cultured
  • DNA / metabolism
  • DNA-Binding Proteins / metabolism
  • DNA-Binding Proteins / physiology*
  • Liver / cytology
  • Liver / metabolism
  • Liver / physiology*
  • Molecular Sequence Data
  • Mutation / physiology
  • Promoter Regions, Genetic / physiology*
  • Protein Processing, Post-Translational / physiology
  • Proto-Oncogene Proteins c-fos / metabolism
  • Proto-Oncogene Proteins c-jun / metabolism
  • Rats
  • Rats, Sprague-Dawley
  • Tissue Inhibitor of Metalloproteinase-1 / genetics*
  • Transcription Factor AP-1 / physiology*
  • Transcription Factors / metabolism
  • Transcriptional Activation / physiology


  • DNA-Binding Proteins
  • Etv5 protein, rat
  • Proto-Oncogene Proteins c-fos
  • Proto-Oncogene Proteins c-jun
  • Tissue Inhibitor of Metalloproteinase-1
  • Transcription Factor AP-1
  • Transcription Factors
  • transcription factor PEA3
  • DNA