Membrane-bound progesterone receptor expression in human aortic endothelial cells.

Observational studies demonstrate that estradiol and progesterone affect vasoreactivity. In animal studies, progesterone treatment causes immediate relaxation of precontracted arteries with inhibition of calcium influx in vascular endothelial and smooth muscle cells, suggesting a non-genomic mechanism of action. In this study we investigated the presence of novel membrane-bound progesterone receptors in human aortic endothelial cells and correlated the expression with cell-cycle stage. Western blotting analysis with an antibody directed to the hormone-binding domain of the classic progesterone receptors shows predominant bands at 100 and 60 kD, whereas analysis with an antibody to the DNA-binding region shows only the 100-kD band. In contrast, classic nuclear progesterone receptors B and A are identified at 116 and 94 kD in similarly processed T47D cells. Both novel bands localize to the membrane fraction after differential centrifugation. Plasma membrane-bound progesterone receptor was further shown with immunofluorescent antibody and ligand-binding studies in a small percentage of human aortic endothelial cells. Fluorescent activated cell sorting demonstrated that approximately 8% of the human aortic endothelial cells expressed a plasma membrane progesterone receptor and that a greater percentage of the expressing cells were in the G2/M-phase of the cell cycle. Treatment with progesterone conjugated to BSA did not show any significant cell-cycle changes. Plasma membrane-bound progesterone receptor in vascular endothelial cells may regulate the non-genomic actions of progesterone, and expression of the receptor appears to vary with cell cycle stage.

Cloning and expression of a novel, truncated, progesterone receptor.

Progesterone acts via two specific receptors to affect gene transcription in target tissues. Progesterone receptor (PR) B contains 933 amino acids while PR A is a truncated version lacking the initial 164 amino acids. We have cloned a novel, truncated PR from both human adipose and aortic cDNA libraries. This cDNA encodes a predicted protein of 314 amino acids, termed PR-M. Initiation of transcription of PR-M occurs in intron 3, with the initial exon identical to exon 4 of the genomic PRs, except for a novel 16 amino acid amino-terminal sequence, consistent with a signal peptide. The remainder of PR-M is identical to the genomic PR. Transcript for this protein was identified by RT-PCR in human aortic endothelial cells and T47D breast cancer cells. Expression of PR-M in Sf 9 insect cells results in a 38-kDa protein, demonstrated in human aortic endothelial cells and T47D breast cancer cells. The function of PR-M remains to be determined. The presence of a signal peptide and the lack of a DNA binding region suggests a non-genomic action.