Poster Presentation The Annual Scientific Meeting of the Endocrine Society of Australia and the Society for Reproductive Biology 2012

DNA methylation of progesterone receptor isoforms in term human myometrium. (#276)

Sze Yee Chai 1 , John Fitter 1 , Roger Smith 1 , Gemma Madsen 1
  1. University of Newcastle/Mothers & Babies Research Centre, Newcastle, NSW, Australia

Nuclear progesterone receptor (PR) isoforms, PR-A and PR-B, are transcribed from a single gene regulated by two separate promoters. Both isoforms bind progesterone and regulate distinct cohorts of genes that contain progesterone response elements. Throughout pregnancy, myometrial PR-A expression is relatively low compared to PR-B, which is the predominant isoform expressed. During active labour, the PR-A:PR-B expression ratio is increased and we postulated that this increase in PR-A expression mediates functional progesterone withdrawal. We hypothesised that the independent regulation of PR isoform expression occurs through epigenetic modifications at the two promoters. We have previously shown that histones surrounding the PR-A promoter are trimethylated at the H3K4 site relative to the PR-B promoter, which would promote increased PR-A expression1. An alternative mechanism for down-regulating gene expression is DNA methylation of CpG islands in the promoter region and first exon of target genes. The PR gene contains CpG islands in both PR-A and PR-B promoter regions, and methylation of these CpG islands has been reported in cancers and cell lines. However, no studies have assessed the methylation status of the PR promoters in pregnant human myometrium. The aims of this study were to determine (1) the methylation status of the PR promoters, and (2) if there are labour-associated changes in the methylation status of the two PR promoters, in human myometrium. Genomic DNA extracted from term myometrial samples (not in labour, n=4; in labour, n=4) were subjected to bisulfite conversion. Promoter regions of PR were amplified and cloned into pGEM-T vector. For each sample, plasmids from ten colonies were purified and sequenced. Our data showed that at term, both PR-A and PR-B promoters are hypomethylated in the myometrium with no labour-associated changes observed [PR-A (not in labour=98%, in labour=96.8% unmethylated); PR-B (not in labour=96.4%, in labour=98.6% unmethylated)]. This suggests that DNA methylation at these regions may not be responsible for the expression changes observed with labour onset.

  1. Chai SY, Smith R, Zakar T, Mitchell C, Madsen G. Term myometrium is characterized by increased activating epigenetic modifications at the progesterone receptor-A promoter. Mol Hum Reprod 2012