Multiple Signal Transduction Pathways through Two Prostaglandin E Receptor EP3 Subtype Isoforms Expressed in Human Uterus

• Published in: The journal of clinical endocrinology and metabolism 2000-11, Vol.85 (11), p.4315-4322
• Main Authors: Kotani, Masato, Tanaka, Issei, Ogawa, Yoshihiro, Suganami, Takayoshi, Matsumoto, Tsunekazu, Muro, Seiji, Yamamoto, Yuji, Sugawara, Akira, Yoshimasa, Yasunao, Sagawa, Norimasa, Narumiya, Shuh, Nakao, Kazuwa
• Format: Article
• Language: English
• Subjects: Alprostadil - analogs & derivatives; Alprostadil - pharmacology; Amino Acid Sequence; Animals; Biological and medical sciences; Cell Membrane - physiology; CHO Cells; Colforsin - pharmacology; Cricetinae; Cyclic AMP - metabolism; Female; Fundamental and applied biological sciences. Psychology; Hormone metabolism and regulation; Humans; Mammalian female genital system; Molecular Sequence Data; Protein Isoforms - genetics; Protein Isoforms - physiology; Receptors, Prostaglandin E - agonists; Receptors, Prostaglandin E - genetics; Receptors, Prostaglandin E - physiology; Receptors, Prostaglandin E, EP3 Subtype; Recombinant Proteins - chemistry; Recombinant Proteins - metabolism; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger - genetics; Sequence Alignment; Signal Transduction - physiology; Transcription, Genetic; Transfection; Uterus - physiology; Vertebrates: reproduction
• ISSN: 0021-972X; 1945-7197
• DOI: 10.1210/jcem.85.11.6989

PGE2 is known to induce uterine contraction by increasing intracellular Ca2+. In the present study, to investigate other functions of PGE2 in human uterus, two EP3 isoforms were isolated by the RT-PCR method using human uterus polyadenylated ribonucleic acid (RNA). These EP3 isoforms, named EP3-V and EP3-VI, are composed of 402 and 393 amino acid residues, respectively, which are unique compared with EP3 isoforms of other species. Their N-terminal 359 amino acid residues are identical to those of previously reported human EP3 isoforms, whereas the two isoforms contained a novel amino acid sequence in their C-terminal tails. The dissociation constant values of EP3-V and EP3-VI for PGE2 were 3.9 and 1.4 nmol/L, respectively, which were consistent with those of previously reported EP3 isoforms. Signaling experiments revealed that M&B28767, an EP3 agonist, not only inhibited forskolin-induced cAMP concentrations, but also activated mitogen-activated protein kinase in Chinese hamster ovary cells stably expressing EP3-V and EP3-VI. These responses were abolished by treatment with pertussis toxin. In addition, M&B28767 increased cAMP concentrations in EP3-VI-expressing cells, whereas it did not in EP3-V-expressing cells. M&B28767 did not stimulate phosphoinositide turnover in EP3-V- or EP3-VI-expressing cells. EP3-V and EP3-VI messenger RNAs (mRNAs) were detected abundantly in human uterus, whereas weak, but substantial, bands were detected in the lung and kidney in RT-PCR specific for each mRNA. In situ hybridization revealed EP3-V and EP3-VI mRNAs in the human myometrium, but not in the endometrium. The present study suggests that EP3-V and EP3-VI are possibly involved in the proliferation of cells in human myometrium.