DNase I-hypersensitive sites enhance alpha1(I) collagen gene expression in hepatic stellate cells.
Liver fibrosis is characterized by a dramatic increase in the expression of type I collagen. Several deoxyribonuclease (DNase) I-hypersensitive sites (HS) have been located in the distal 5'-flanking region of the alpha1(I) collagen gene that are specific to collagen-producing cells. To assess the role of the DNase I-HS in regulating alpha1(I) collagen gene expression in hepatic stellate cells (HSCs), 3 transgenic mouse lines expressing collagen-alpha1(I) reporter genes were used (Krempen et al. Gene Expr 1999;8:151-163). The pCol9GFP transgene contains the collagen gene promoter (-3122 to +111) linked to the green fluorescent protein (GFP) reporter gene. The pCol9GFP-HS4,5 transgene contains HS4,5 and pColGFP-HS8,9 contains HS8,9 positioned upstream of the collagen promoter in pCol9GFP. HSCs isolated from transgenic mice containing pCol9GFPHS4,5 and pColGFP-HS8,9 showed earlier and higher GFP expression patterns than HSCs isolated from pCol9GFP mice. HSCs from pCol9GFP-HS4,5 showed the highest levels of GFP expression and culture-induced expression correlated with induction of the endogenous alpha1(I) collagen gene. After CCl(4) administration, pCol9GFP-HS4,5 mice showed increased GFP expression compared with pCol9GFP mice in both whole liver extracts and isolated HSCs. Several sites for DNA-protein interactions in both HS4 and HS5 were identified that included a binding site for activator protein 1. In conclusion, DNase I-HS4,5 enhance expression of the alpha1(I) collagen gene promoter in HSCs both in vitro and in vivo after a fibrogenic stimulus. The collagen-GFP transgenic mice provide a convenient and reliable model system to investigate the molecular mechanisms controlling increased collagen expression during fibrosis.
Yata
Yutaka
Y
Department of Medicine, The University of North Carolina at Chapel Hill, 27599, USA.
Scanga
Andrew
A
Gillan
Andrea
A
Yang
Liu
L
Reif
Shimon
S
Breindl
Michael
M
Brenner
David A
DA
Rippe
Richard A
RA
D000818:Animals; D001483:Base Sequence / Q000235:genetics; D001665:Binding Sites / Q000235:genetics; D002251:Carbon Tetrachloride; D002478:Cells, Cultured; D024042:Collagen Type I / Q000235:genetics*; D004247:DNA / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000502:physiology*; D015870:Gene Expression / Q000502:physiology*; D015971:Gene Expression Regulation, Enzymologic; D022781:Hepatocytes / Q000502:physiology*; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D008103:Liver Cirrhosis / Q000139:chemically induced / Q000235:genetics; D051379:Mice; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic / Q000235:genetics; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011506:Proteins / Q000378:metabolism; D019076:Transgenes / Q000502:physiology
D024042:Collagen Type I; D011506:Proteins; D004247:DNA; D002251:Carbon Tetrachloride; D003850:Deoxyribonuclease I