Decreased DNA Methylations at the Progesterone Receptor Promoter A Induce Functional Progesterone Withdrawal in Human Parturition

Abstract

The functional interaction of progesterone receptor (PR) isoforms PRA and PRB regulates myometrial transition from the resting state to excitation-contraction to initiate parturition. However, the regulatory mechanisms responsible for maintenance and functional alteration of the PRA and PRB expression levels during human pregnancy and term labor, respectively, remain unknown. Therefore, this study was designed to investigate whether and how epigenetic DNA modifications, specifically methylations, at the PRs' promoter regions contribute to the differential expression of PRA and PRB in laboring term myometrium of humans. Comparative analysis of PRA and PRB messenger RNA (mRNA) expression levels and accompanying changes in their promoters' methylation status was carried out using human myometrial samples from women undergoing singleton, term deliveries by cesarean section, either in the absence of labor (designated as NIL for not-in-labor) or in active labor (designated as IL for in labor). The PRA gene expression was shown to be elevated significantly during labor, while PRB gene expression was unaltered, and this differential expression was accompanied by decreased DNA methylation at the PRA promoter and not at the PRB promoter. In addition, labor-related decreased mRNA expression of the DNA methyltransferase (DNMT) family members DNMT1 and DNMT3a was found, however whether the increased expression of DNMTs directly supports the functional withdrawal of progesterone needs further investigation. Collectively, these data indicate that DNA methylation might represent an important epigenetic mechanism of labor-related differential expression of PRs, thereby mediating the biological process of functional PR withdrawal at term for parturition.

Keywords: DNA methylation; DNMT; labor; myometrium; progesterone receptors.

Figure 1.

The CpG island and CpG amplification primer fragment in the promoter of PRA and PRB genes. Schematic diagram showing the transcription start site of PRA and PRB genes. The CpG island location and the CpG amplification primer fragment in the promoter region of PRA and PRB genes were indicated, respectively. PR indicates progesterone receptor.

Figure 2.

Myometrium mRNA expression of PRA and PRB prior to and after labor onset. Panel A, relative mRNA abundance of PRA and PRB in human myometrial samples collected prior to labor onset (NIL, n = 25) or after labor onset (IL, n = 25), expressed relative to the β-actin reference gene. Panel B, PRA-PRB mRNA expression ratio prior to and after labor onset (n = 25). Data are presented as mean ± SD. *P < .05 versus the corresponding NIL group; **P < .01 versus the NIL group. mRNA indicates messenger RNA; PR, progesterone receptor; NIL, not in labor; IL, in labor; SD, standard deviation.

Figure 3.

Protein expression of PRs during labor. PR total (left) and PRB protein (right) expression (brown) is shown in NIL and IL myometrium tissues. PR indicates progesterone receptor; NIL, not in labor; IL, in labor.

Figure 4.

DNA methylation of PRA and PRB during labor. Methylation status of PRA (left) and PRB (right) is shown. The 2 methylated CpG sites in PRB were summed and presented as total CpG islands for the gene region. Data are plotted as mean (n = 6) ± SD. **P < .01 versus the NIL group. SD indicates standard deviation; PR, progesterone receptor; NIL, not in labor.

Figure 5.

The mRNA expression of DNMTs in myometrium during labor. The relative mRNA abundance of DNMT1 (right) and DNMT3a and DNMT3b (left) in human myometrial samples is shown. Relative expression was determined by comparison to the β-actin reference gene. Data are plotted as mean (n = 25) ± SD. *P < .05 versus the corresponding NIL group. mRNA indicates messenger RNA; DNMT, DNA methyltransferase; SD, standard deviation.