Abstract Our recent study demonstrated that upregulation of Rad, a monomeric G protein, is a novel regulatory mechanism inhibiting Ca V 1.2 channel currents in uterine artery hemodynamic adaptation to pregnancy in a sheep model. In the present study, we test the hypothesis that chronic hypoxia during gestation downregulates Rad gene expression via promotor hypermethylation and impairs Rad‐mediated inhibition of Ca V 1.2 channel activity in uterine arteries. We showed that uterine artery Ca V 1.2 current density and vascular resistance were significantly increased in pregnant ewes acclimatized to high altitude hypoxia, as compared to normoxic control animals. RNA‐seq analysis revealed a highly significant downregulation of Rad gene in uterine arteries that was validated by qRT‐PCR and Western blots. We found that the ovine Rad gene promoter contains a CG island adjacent to the transcription start site and it was hypermethylated in uterine arteries of pregnant ewes exposed to chronic hypoxia, resulting in a decrease in the promoter activity. Knockdown of ten‐eleven translocation methylcytosine dioxygenase 2 (TET2) recapitulated the effect of hypoxia, increasing Rad promoter methylation, decreasing Rad gene expression, and upregulating Ca V 1.2 activity in uterine arteries. Rad overexpression in vivo in ovine uterine artery reversed gestational hypoxia‐induced increases in both Ca V 1.2 activity and myogenic tone, albeit with no significant effect observed on uterine artery vascular resistance. Together, our study reveals a novel mechanism of hypermethylation in hypoxia‐mediated Rad downregulation and Ca V 1.2 hyperactivity of uterine arteries during pregnancy, providing new insights into the understanding of uterine hemodynamic maladaptation in maternal complications associated with gestational hypoxia. image Key points Chronic gestational hypoxia increases uterine artery Ca V 1.2 current density and vascular resistance in pregnant ewes. RNA‐seq analysis reveals a highly significant downregulation of Rad gene in uterine arteries of hypoxic pregnant animals. Rad gene promoter is hypermethylated in uterine arteries of hypoxic pregnant animals and increased methylation results in a decrease in Rad promoter activity. Knockdown of ten‐eleven translocation methylcytosine dioxygenase 2 (TET2) recapitulates the effect of hypoxia by increasing Rad promoter hypermethylation, decreasing Rad gene expression, and upregulating Ca V 1.2 activity in uterine arteries. Rad overexpression in vivo negates gestational hypoxia‐induced increases in uterine artery Ca V 1.2 activity and myogenic tone.
Hu et al. (Sat,) studied this question.