CCAAT binding transcription factor binds and regulates human COL1A1 promoter activity in human dermal fibroblasts: demonstration of increased binding in systemic sclerosis fibroblasts.

Abstract

OBJECTIVE:

To determine the binding factors that interact with the proximal promoter region of the human type I collagen gene, COL1A1, and to examine their involvement in its transcriptional regulation in normal and systemic sclerosis (SSc) dermal fibroblasts.

METHODS:

Nuclear extracts from dermal fibroblasts from 4 patients with SSc and 4 age- and sex-matched control individuals were examined by electrophoresis mobility shift assays with a COL1A1 promoter fragment encompassing nucleotides -174 to -50 bp. Supershift assays with antibodies specific to various transcription factors, and competition experiments using consensus, wild-type, or mutated oligonucleotides corresponding to their specific binding sites, were performed. The effects of specific oligonucleotides as "intracellular competitors" were examined by transient transfection experiments in SSc fibroblasts using a COL1A1 construct containing -174 bp of the promoter.

RESULTS:

The findings demonstrate that the CCAAT binding transcription factor (CBF) binds the proximal CCAAT box located at -100 to -96 bp, but not the distal CCAAT box at -125 to -121 bp, of the human COL1A1 promoter in both SSc and normal fibroblasts. CBF binding activity was 3-5-fold higher in the SSc fibroblasts. Moreover, the promoter activity of the -174-bp COL1A1 construct was decreased by up to 50% when specific oligonucleotides were used as "intracellular competitors." In addition, Sp1 and Sp3 were other transcription factors found to be involved in the formation of the DNA-protein complexes within this region of the COL1A1 promoter.

CONCLUSION:

These results indicate that the transcription factor CBF binds the human COL1A1 proximal promoter region in human dermal fibroblasts, and its binding activity is higher in SSc fibroblasts.

• Authors: B Saitta, S Gaidarova, L Cicchillitti, S A Jimenez
• Journal: Arthritis and rheumatism (Arthritis Rheum) Vol. 43 Issue 10 Pg. 2219-29 (Oct 2000) ISSN: 0004-3591 [Print] United States
• PMID: 11037881 (Publication Type: Journal Article, Research Support, U.S. Gov't, P.H.S.)

Chemical References

• CCAAT-Binding Factor
• DNA-Binding Proteins
• Oligonucleotide Probes
• SP3 protein, human
• Sp1 Transcription Factor
• Transcription Factors
• Sp3 Transcription Factor
• Collagen

Topics

• Base Sequence
• Binding, Competitive (drug effects)
• CCAAT-Binding Factor (metabolism, pharmacology)
• Cells, Cultured
• Collagen (genetics)
• DNA-Binding Proteins (pharmacology)
• Fibroblasts (metabolism)
• Gene Expression Regulation (drug effects)
• Humans
• Oligonucleotide Probes
• Promoter Regions, Genetic (drug effects, genetics)
• Scleroderma, Systemic (metabolism, pathology)
• Skin (pathology)
• Sp1 Transcription Factor (pharmacology)
• Sp3 Transcription Factor
• Transcription Factors (metabolism, pharmacology)