| Co-Investigator(Kenkyū-buntansha) |
NATA Koji Tohoku University, Graduate School of Medicine, Research Associate, 大学院医学系研究科, 助手 (90202233)
NOGUCHI Naoya Tohoku University, Graduate School of Medicine, Research Associate, 大学院医学系研究科, 助手 (20333792)
岡本 宏 東北大学, 大学院・医学系研究科, 教授 (60025632)
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| Budget Amount *help |
¥14,700,000 (Direct Cost: ¥14,700,000)
Fiscal Year 2006: ¥5,300,000 (Direct Cost: ¥5,300,000)
Fiscal Year 2005: ¥9,400,000 (Direct Cost: ¥9,400,000)
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| Research Abstract |
Intracellular Ca^<2+> mobilization in a variety of cells is known to be mediated by two major systems, the inositol 1,4,5-trisphosphate and cyclic ADP-ribose (cADPR) systems, and in normal islets, it has been proposed to be mediated by the cADPR-dependent system (the CD38-cADPR signal system). That is, cADPR generated by glucose stimulation acts as a second messenger for Ca^<2+> release from the endoplasmic reticulum for insulin secretion. The ryanodine receptor (RyR) has been postulated to be an intracellular Ca^<2+> release channel for cADPR in a wide variety of tissues including pancreatic islets. To date, three different RyRs encoded by separate genes have been identified, namely the skeletal-type RyR1, the cardiac-type RyR2 and the brain-type RyR3. RyR1 and RyR2 function as Ca^<2+>-induced Ca^<2+> release (CICR) channels for the contraction of skeletal and cardiac muscles, respectively, and RyR3 was first isolated from brain. It has been demonstrated that cADPR binds to FK506-bind
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ing protein 12.6 (FKBP 12.6) on rat islet ryanodine receptor and that the binding of cADPR to FKBP12.6 frees RyR2 from FKBP12.6, causing it to release Ca^<2+>. These 3 channels are expressed in many tissues including skeletal muscle, heart, brain, and pancreatic islets. Recently, a variety of splicing isoforms of RyRs were isolated, and some of them showed differences in terms of the channel function properties. In the present study, by screening a rat islet cDNA library, we isolated a novel RyR cDNA (the islet-type RyR), which is generated from the RyR2 gene by alternative splicing of exons 4 and 75. The alternatively spliced exons are found in the mammalian RyR2 gene but not in the mammalian RyR1, RyR3, nor in the Drosophila RyR gene, suggesting that these exons were acquired in the evolution of the RyR2 gene from an ancestral RyR gene. When the expression vectors for the islet-type and the authentic RyRs were transfected into HEK293 cells, the islet-type RyR2 as well as the authentic one showed high affinity [^3H]ryanodine binding. Intracellular Ca^<2+> release in the islet-type RyR2-transfected cells was enhanced in the presence of cADPR but not in the authentic RyR2-transfected cells. The islet-type RyR2 mRNA was expressed in a variety of tissues such as in pancreatic islets, cerebrum, and cerebellum, whereas the authentic RyR2 mRNA was predominantly expressed in heart and aorta. These results strongly suggest that the islet-type RyR2 is an intracellular target for cADPR signaling. Less
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