Critical role of the N‐terminal cyclic AMP‐binding domain of Epac2 in its subcellular localization and function |
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Authors: | Manabu Niimura Takashi Miki Tadao Shibasaki Wakako Fujimoto Toshihiko Iwanaga Susumu Seino |
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Institution: | 1. Division of Cellular and Molecular Medicine, Department of Physiology and Cell Biology, Kobe University Graduate School of Medicine, Kobe, Hyogo, Japan;2. Department of Autonomic Physiology (C3), Chiba University Graduate School of Medicine, Chiba, Chiba, Japan;3. Laboratory of Histology and Cytology, Graduate School of Medicine, Hokkaido University, Sapporo, Hokkaido, Japan;4. Division of Diabetes, Metabolism and Endocrinology, Department of Internal Medicine, Kobe University Graduate School of Medicine, Kobe, Hyogo, Japan;5. Core Research for Evolutional Science and Technology (CREST), Japan Science and Technology Agency, Kawaguchi, Saitama, Japan |
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Abstract: | cAMP is a well‐known regulator of exocytosis, and cAMP‐GEFII (Epac2) is involved in the potentiation of cAMP‐dependent, PKA‐independent regulated exocytosis in secretory cells. However, the mechanisms of its action are not fully understood. In the course of our study of Epac2 knockout mice, we identified a novel splicing variant of Epac2, which we designate Epac2B, while renaming the previously identified Epac2 Epac2A. Epac2B, which lacks the first cAMP‐binding domain A in the N‐terminus but has the second cAMP‐binding domain B of Epac2A, possesses GEF activity towards Rap1, as was found for Epac2A. Immunocytochemical analysis revealed that exogenously introduced Epac2A into insulin‐secreting MIN6 cells was localized near the plasma membrane, while Epac2B was found primarily in the cytoplasm. Interestingly, cAMP‐binding domain A alone introduced into MIN6 cells was also localized near the plasma membrane. In MIN6 cells, Epac2A was involved in triggering hormone secretion by stimulation with 5.6 mM glucose plus 1 mM 8‐Bromo‐cAMP, but Epac2B was not. The addition of a membrane‐targeting signal to the N‐terminus of Epac2B was able to mimic the effect of Epac2A on hormone secretion. Thus, the present study indicates that the N‐terminal cAMP‐binding domain A of Epac2A plays a critical role in determining its subcellular localization and potentiating insulin secretion by cAMP. J. Cell. Physiol. 219: 652–658, 2009. © 2009 Wiley‐Liss, Inc. |
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