EP3 receptor isoforms are differentially expressed in subpopulations of primate granulosa cells and couple to unique G-proteins

Abstract

Prostaglandin E2 produced within the ovarian follicle is necessary for ovulation. Prostaglandin E2 is recognized by four distinct G-protein coupled receptors. Among them, PTGER3 (also known as EP3) is unique in that mRNA splicing generates multiple isoforms. Each isoform has a distinct amino acid composition in the C-terminal region, which is involved in G-protein coupling. To determine if monkey EP3 isoforms couple to different G-proteins, each EP3 isoform was expressed in Chinese hamster ovary (CHO) cells, and intracellular signals were examined after stimulation with the EP3 agonist sulprostone. Stimulation of EP3 isoform 5 (EP3-5) reduced cyclic adenosine monophosphate (cAMP) in a pertussis toxin-sensitive manner, indicating involvement of Gαi. Stimulation of EP3-9 increased cAMP, which was reduced by the general G-protein inhibitor GDP-β-S, and also increased intracellular calcium, which was reduced by pertussis toxin and GDP-β-S. So, EP3-9 likely couples to both Gαs and a pertussis toxin-sensitive G-protein to regulate intracellular signals. Stimulation of EP3-14 increased cAMP, which was further increased by pertussis toxin, so EP3-14 likely regulates cAMP via multiple G-proteins. Granulosa cell expression of all EP3 isoforms increased in response to an ovulatory dose of hCG. Two EP3 isoforms were differentially expressed in functional subpopulations of granulosa cells. EP3-5 was low in granulosa cells at the follicle apex while EP3-9 was high in cumulus granulosa cells. Differential expression of EP3 isoforms may yield different intracellular responses to prostaglandin E2 in granulosa cell subpopulations, contributing to the different roles played by granulosa cell subpopulations in the process of ovulation.

Prostaglandin E2 promotes human cholangiocarcinoma cell proliferation, migration and invasion through the upregulation of β-catenin expression via EP3-4 receptor

“…These unique sequences in the C-terminal domains may allow each EP3 isoform to localize to different areas within the cell and trigger different intracellular signaling pathways in response to PGE2. Evidence of different signal transduction pathways and the regulation of gene expression among different EP3 receptor isoforms has also been previously demonstrated (12,17). There are EP1, EP2, EP4 and four EP3 isoforms receptors involved in cholangiocarcinoma progression, and some receptors play a positive role while others do not.…”

“…Five isoforms of the human EP3 receptors: EP3-4, EP3-5, EP3-6, EP3-7 and EP3-8 have been identified. Evidence of different signal transduction pathways and the regulation of gene expression among different EP receptors has also been demonstrated in a number of studies (17). Thus, the specific target of PGE2 in regulating cancer cell proliferation via EP receptors have yet to be clarified.…”

Mapping PTGERs to the Ovulatory Follicle: Regional Responses to the Ovulatory PGE2 Signal

“…PTGER3 is unique among PTGERs in that multiple isoforms are formed by alternative RNA splicing, resulting in proteins of 40–45 kDa [ 81 – 91 ]. PTGER3 isoforms have essentially identical amino acid sequences in the ligand-binding region and, therefore, possess very similar ligand-binding properties.…”

“…Cumulus expression of PTGER3 has been reported for mouse, cow, and monkey [ 138 , 142 , 144 ], but increased PTGER3 mRNA has been reported after the ovulatory gonadotropin surge only in primates [ 144 ]. In monkey cumulus, the PTGER3 isoform 9 was the predominant PTGER3 isoform detected, with mRNA levels elevated 100-fold after the ovulatory gonadotropin surge [ 91 ]. The presence of receptor protein for PTGER2 [ 54 , 138 , 143 ], PTGER3 [ 138 , 144 ], and PTGER4 [ 138 , 144 ] has been confirmed for cumulus from several species.…”

“…Mural granulosa cell expression of PTGER2 and PTGER4 was shown to increase in response to the ovulatory gonadotropin surge in horse and monkey [ 143 , 144 ]. Additional studies identified five distinct PTGER3 isoforms with increased mRNA levels after the ovulatory gonadotropin surge in monkey mural granulosa cells [ 91 ]. The ovulatory gonadotropin surge also led to significant increases in granulosa cell levels of PTGER1, PTGER3, and PTGER4 proteins in monkey [ 144 ] and PTGER2 protein in horse [ 143 ].…”