CpG methylation represses the activity of the rat prolactin promoter in rat GH3 pituitary cell lines

Molecular and Cellular Endocrinology
V M NgôD Gourdji

Abstract

In the present report, we have investigated the role of DNA methylation on the binding and trans-acting properties of transcription factors involved in the regulation of the rat prolactin (rPRL) gene, specifically Pit-1. To this aim we took advantage of a model system composed of three GH3 rat pituitary tumor cell lines that greatly differed in the extent of rPRL gene methylation and in the level of rPRL gene expression. Northern blot analyses indicated that identical species of Pit-1 mRNA were present to similar extent in the three GH3 cell lines. Electrophoretic mobility shift assays further demonstrated that Pit-1 was present in nuclear extracts and displayed equal affinities to bind the 1P responsive element encompassing the -65 to -38 region of the rPRL promoter, whatever the GH3 cell line tested. These data suggested that differential expression of the rPRL gene among cell lines did not result from variable amounts of Pit-1. By combining in vitro methylation and transient transfection experiments with a rPRL promoter-driven CAT construct, we showed that extensive methylation at CpG sites abolished the expression of the reporter gene. Furthermore, in vivo competition assays demonstrated that CpG methylation inhibited gene ...Continue Reading

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Jan 3, 2001·Brain Research·J H BoatrightD E Borst
Feb 14, 2002·Differentiation; Research in Biological Diversity·Kunio Shiota, Ryuzo Yanagimachi
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Apr 5, 2018·Frontiers in Pharmacology·Yu WangLizhong Du
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