All-trans-Retinoic Acid Inhibition of Proα1(I) Collagen Gene Expression in Fetal Rat Skin Fibroblasts: Identification of a Retinoic Acid Response Element in the Proα1(I) Collagen Gene

Abstract

The current study was undertaken to determine the mechanism by which the retinoid all-trans-retinoic acid regulates pro alpha1(I) collagen gene expression in fetal rat skin fibroblasts. FRS fibroblasts were stably transfected with the ColCat3.6 plasmid, which contains a portion of the 5' flanking region of the rat pro alpha1(I) collagen gene linked to a reporter gene, chloramphenicol acetyltransferase. The effect of t-RA on CAT activity was determined as a function of concentration and incubation time. Maximal inhibition of CAT activity by t-RA occurred at 10(-8) M after 48 h of treatment. Transforming growth factor-beta1 did not block the inhibitory effect of t-RA on CAT activity. Computer sequence analysis of the 3.6-kb DNA fragment that contains the promoter for the rat pro alpha1(I) collagen gene identified a direct repeat RARE sequence composed of one diverse (5'-AGTAGA-3') and one idealized (5'-GGGTCA-3') half site located at positions -1345 and -1335, respectively. Two nuclear retinoid receptors that were expressed in bacteria, retinoic acid receptor-gamma and retinoid X receptor-alpha, were found to bind specifically to a double-stranded oligonucleotide containing the RARE in gel mobility shift assays. Mutation of the idealized half-site eliminated the binding of receptor proteins to the oligonucleotide. Gel mobility shift assays using nuclear protein extracts prepared from t-RA-treated FRS fibroblasts showed that binding to the oligonucleotide containing the RARE was decreased from control values. The same assays performed with the mutated oligonucleotide resulted in only slight binding. These studies indicate that t-RA downregulates the promoter activity of the rat pro alpha1(I) collagen gene by decreasing the binding of nuclear protein to the RARE sequence in the 5' flanking region of the gene.

Coordinated gene expression during gilthead sea bream skeletogenesis and its disruption by nutritional hypervitaminosis A

“…Collagen fibres comprise 90% of the ECM proteins in skeletal tissues and confer most of their physical properties [84]. The present ontogeny of the COL1A1 expression was in agreement with previous results found in European sea bass, where COL1A1 was highly expressed from 31 dph onwards [14].…”

“…Different results from the effects of RA on the expression of COL1A1 are reported in the literature [6], which suggests an indirect regulation of COL1A1 by RA. However, larvae from the 1.5×VA and 10×VA groups showed lower expression of COL1A1 at 60 dph, which seems to be attributed to the lower expression of RARG detected in those animals, since this RAR binds specifically to the RARE of the COL1A1 promoter [84]. …”

The effect of topical tretinoin on tissue strength and skin components in a murine incisional wound model

“…28 This inhibitory effect is consistent with studies which have shown that tretinoin effectively reduces the proliferation of fibroblasts as well as decreases fibroblast collagen synthesis. 29,30 In contrast, Basak et al 13 reported enhancement of collagen production, angiogenesis and granulation tissue formation under tretinoin treatment.…”

Retinoids and nephron mass control

“…This change involves a shift from type I and type III collagens to type IV collagen. Recently, it has been shown that RA downregulates the promoter activity of the rat pro-α 1 -collagen type I [49]. Applied to the developing kidney, this may help differentiation of mesenchymal cells into tubular structures.…”