Hypermethylation of the hTERT promoter inhibits the expression of telomerase activity in normal oral fibroblasts and senescent normal oral keratinocytes.

Abstract

Telomerase activity in human cells closely correlates with the expression of its catalytic subunit, telomerase reverse transcriptase (hTERT). Previously, we reported the lack of telomerase activity in normal human oral fibroblasts (NHOF) and the diminution of telomerase activity during senescence in normal human oral keratinocytes (NHOK). To investigate the underlying mechanisms of telomerase regulation in both cell types, we analysed the expression, promoter activity, and methylation status of the hTERT gene. The expression of hTERT mRNA diminished in senescent NHOK, but was not detected in NHOF at any stage of replication. An exogenous hTERT promoter was active in NHOF and in senescing NHOK, indicating that the lack of hTERT gene expression resulted from alteration of the endogenous hTERT promoter. Since methylation is involved in the silencing of numerous genes, we carried out DNA methylation assays. The assay revealed that the hTERT promoter was hypermethylated in NHOF and was gradually methylated during senescence in NHOK. Treatment of NHOF and senescent NHOK with the demethylating agent 5-aza-2′-deoxycytidine restored the expression of endogenous hTERT mRNA. Our results suggest that hypermethylation of the hTERT promoter plays a critical role in the negative regulation of telomerase activity in normal human oral cells.