title: NF-kappaB inhibits expression of the alpha1(I) collagen gene.
abstract: Fibrosis results from an increase in the synthesis and deposition of type I collagen. Fibrosis is frequently associated with inflammation, which is accompanied by increased levels of tumor necrosis factor-alpha (TNFalpha) and activation of the transcription factor NF-kappaB. However, several agents known to activate NF-kappaB, such as phorbol 12-myristate 13-acetate (PMA) and TNFalpha, result in decreased expression of type I collagen. Therefore, we directly examined the effects of NF-kappaB on alpha1(I) collagen gene expression in two collagen-producing cells, NIH 3T3 fibroblasts and hepatic stellate cells (HSCs). Transient transfections of NIH 3T3 cells or HSCs using NF-kappaB p50, p65, and c-Rel expression plasmids with collagen reporter gene plasmids demonstrated a strong inhibitory effect on transcription of the collagen gene promoter. Dose-response curves showed that p65 was a stronger inhibitor of collagen gene expression than was NF-kappaB p50 or c-Rel (maximum inhibition 90%). Transient transfections with reporter gene plasmids containing one or two Spl binding sites demonstrated similar inhibitory effects of NF-kappaB p65 on the activity of these reporter genes, suggesting that the inhibitory effects of NF-kappaB p65 are mediated through the critical Spl binding sites in the alpha1(I) collagen gene promoter. Cotransfection experiments using either a super-repressor I[ke]B or Spl partially blocked the inhibitory effects of p65 on collagen reporter gene activity. Coimmunoprecipitation experiments demonstrated that NF-kappaB and Spl do interact in vivo. Nuclear run-on assays showed that NF-kappaB p65 inhibited transcription of the endogenous alpha1(I) collagen gene. Together, these results demonstrate that NF-kappaB decreases transcription of the alpha1(I) collagen gene.
authors.affiliation: Department of Medicine, The University of North Carolina, Chapel Hill 27955-7038, USA. rarippe@med.unc.edu
authors.forename: R A
authors.initials: RA
authors.lastname: Rippe
authors.forename: L W
authors.initials: LW
authors.lastname: Schrum
authors.forename: B
authors.initials: B
authors.lastname: Stefanovic
authors.forename: J A
authors.initials: JA
authors.lastname: Solís-Herruzo
authors.forename: D A
authors.initials: DA
authors.lastname: Brenner
mesh_terms: D016475:3T3 Cells; D000818:Animals; D001665:Binding Sites; D003094:Collagen / Q000096:biosynthesis / Q000235:genetics*; D016384:Consensus Sequence; D005355:Fibrosis; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D020672:I-kappa B Proteins / Q000502:physiology; D008099:Liver / Q000166:cytology; D008103:Liver Cirrhosis / Q000235:genetics; D046911:Macromolecular Substances; D008297:Male; D051379:Mice; D016328:NF-kappa B / Q000235:genetics / Q000502:physiology*; D052002:NF-kappa B p50 Subunit; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D020033:Protein Isoforms / Q000096:biosynthesis / Q000235:genetics* / Q000502:physiology; D020569:Proto-Oncogene Proteins c-rel / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D017207:Rats, Sprague-Dawley; D016329:Sp1 Transcription Factor / Q000378:metabolism; D051996:Transcription Factor RelA; D014162:Transfection
chemical_list: D020672:I-kappa B Proteins; D046911:Macromolecular Substances; D016328:NF-kappa B; D052002:NF-kappa B p50 Subunit; D020033:Protein Isoforms; D020569:Proto-Oncogene Proteins c-rel; D012333:RNA, Messenger; D016329:Sp1 Transcription Factor; D051996:Transcription Factor RelA; D003094:Collagen