CLONING OF HISTAMINE H1-RECEPTOR cDNA
| Project/Area Number |
03670102
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
General pharmacology
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| Research Institution | OSAKA UNIVERSITY |
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Principal Investigator |
FUKUI Hiroyuki OSAKA UNIVERSITY, FACULTY OF MEDICINE, DEPARTMENT OF PHARMACOLOGY II, 医学部, 助教授 (90112052)
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| Co-Investigator(Kenkyū-buntansha) |
ITO Seiji OSAKA BIOSCIENCE INSTITUTE, DEPARTMENT OF CELL BIOLOGY, 副部長 (80201325)
HORIO Yoshiyuki OSAKA UNIVERSITY, FACULTY OF MEDICINE, DEPARTMENT OF PHARMACOLOGY II, 医学部, 助手 (30181530)
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| Project Period (FY) |
1991 – 1992
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| Project Status |
Completed (Fiscal Year 1992)
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| Budget Amount *help |
¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1992: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1991: ¥1,300,000 (Direct Cost: ¥1,300,000)
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| Keywords | HISTAMINE / HISTAMINE RECEPTOR / XENOPUS OOCYTE / CLONING / G PROTEIN-COUPLED RECEPTOR / DEBRISOQUINE / [^3H]MEPYRAMINE BINDING ASSAY / [^3H]メピラミン / プロモーター領域 / デブリソキン水酸化酵素 / ヒスタミンH_1受容体 / クロ-ニング / アフリカツメガエル / ロドプシンファミリ- / Ca^<++>動員受容体 / 副腎髄質 / アフィニティラベル |
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| Research Abstract |
(a) Histamine H1 receptors in bovine adrenal medulla were characterized by [^3H]mepyramine binding assay and by using an affinity labeling reagent (1). When mRNA extracted from bovine adrenal medulla was injected into Xenopus oocytes, expression of H1 receptors were electrophysiologically detected (2). A cDNA library was built from mRNA of bovine adrenal medulla using an expression vector, lambdaZAPII. A histamine H1 receptor cDNA clone was isolated from 65 pools of the library containing 20,000 clones by a combination of cloning in an in vitro transcription of mRNA and electrophysiological assay of H1 receptors expressed in Xenopus oocytes which were injected with mRNA by in vitro transcription (3). The H1 receptor cDNA encoded 491 amino acid residues with molecular weight of 55,954 and seven hydrophobic amino acid sequences which are characteristic to G protein-coupled receptors. Expressed H1 receptors in mammalian cells using the H1 receptor cDNA showed common characteristics of H1
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receptors. Antidepressants, neuroleptics and antiserotonins which have affinities to H1 receptors also showed affinities to the H1 receptor. The expressed H1 receptor mediated histamine-induced inositol phosphates accumulation and Ca^<2+> mobilization.
(b) Rat histamine H1 receptor gene was cloned (4). The clone did not have an intron and encoded 486 amino acid residues. In the promotor region, a glucocorticoid response element and an AP-2 element were observed. A band of H1 receptor mRNA form rat brain was visualized by Northern blot analysis. However, that from intestine was faint and those from lung and heart were not detected, and this results suggested the low sensitivity to histamine.
(c) Another [^3H]mepyramine binding protein than H1 receptors was purified from rat liver and revealed to be a relating protein of debrisoquine 4-hydroxylase, cytochrome P450. The affinity of [^3H]mepyramine binding protein in liver was 1,000 times higher than that of the H1 receptor, and [^3H]mepyramine labeled the H1 receptor in the presence of 10 muM quinine, an inhibitor of debrisoquine 4-hydroxylase. Less
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Report
(3 results)
Research Products
(21 results)
