DNA methylation inhibits transcription of procollagen α2(I) promoters

Abstract

Our previous studies have demonstrated that a 2-[N-(acetoxyacetyl)amino]fluorene-transformed rat epithelial-like cell line, W8, contains a transcriptionally inactive alpha 2(I) gene with a hypermethylated promoter/first-exon region. We have cloned the rat promoter/first-exon region (-211 to +207) from W8 cells and their parent cell line, K16, which expresses alpha 2(I) collagen. There were no sequence differences between the clones from the two cell lines, indicating that a mutation was not responsible for transcriptional inhibition. The alpha 2(I) rat promoters were cloned upstream of the chloramphenicol acetyltransferase gene. Both constructs were equally active in both cell lines, suggesting that trans-activating factors for alpha 2(I) transcription are present in W8 cells. Finally, methylation of plasmids at all CpG sites with SssI methylase completely inhibited transcription using alpha 2(I) promoters, but methylation did not inhibit simian-virus-40 promoter-driven transcription. Certain methylation sites partially inhibit promoter activity. An HhaI methylation site inhibited transcriptional activity of the alpha 2(I) promoter 8-fold, whereas methylation at the HpaII site in the rat alpha 2(I) promoter did not decrease transcriptional activity. This provides further evidence that methylation at specific sites in the collagen alpha 2(I) promoter is responsible for the inactivation of transcription in W8 cells.

Changes in cell cycle and extracellular matrix gene expression during placental development in deer mouse (Peromyscus) hybrids

“…Many of the genes we have identified as misregulated in the Peromyscus hybrids have been shown to be subject to epigenetic regulation, including Timp1, Timp2, Timp3, Procollagen I, S100a6/Calcyclin and Cdkn1c/p57 kip2 (Guenette et al 1992;Wistuba et al 2001;Feng et al 2004;Fan et al 2005;Rehman et al 2005;Chu et al 2006). In addition, Cdkn1c/p57 kip2 is an imprinted gene and is therefore regulated by epigenetic modifications (Hatada et al 1996).…”

“…For example, misregulation of genes involved in the cell cycle (Rb1, Cdkn1c/p57 kip2 , p16), mismatch repair (MlH1), Wnt signalling (SFRP1, SFRP2, SFRP4 and SFRP5), ECM (procollagen-encoding loci), metastasis (Timp3 and s100a6/Calcyclin) and many other pathways leads to defects in the control of proliferation, differentiation and apoptosis. All these loci also exhibit altered DNA methylation at the promoter associated with aberrant transcription (Parker et al 1982;Guenette et al 1992;Baylin and Ohm 2006).…”

DNA methylation represses the murine alpha 1(I) collagen promoter by an indirect mechanism.

“…In the mutant Movl3 mouse strain, transcriptional inactivation of the (xl(I) collagen gene by insertion of a retroviral provirus (10,30) is associated with changes in the methylation pattern of the cellular sequences (16,34). In vitro methylation of the rat a2(I) promoter and the human odl(I) promoter abolishes their transcriptional activity (27,63). The murine al(IV) collagen gene is inactive in undifferentiated F9 teratocarcinoma cells and can be activated by demethylating agents (13,17).…”

Two promoters control the transcription of the human α2(VI) collagen gene

“…A similar discordance of the promoter activity and the mRNA expression was recently reported for the chicken a2(VI) collagen gene (Koller et al, 1991) where this discrepancy was explained by the possibility that DNA regions not included in the promoter constructs are responsible for the cell-specific gene expression. Alternatively, different patterns of DNA methylation may occur in different cell types, and could be responsible for the inhibition of transcription, as was found for the al(1) and the a2(I) collagen genes ( Thompson, et al, 1991;Guenette, et al, 1992).…”