Molecular cloning of a novel gene involved in serotonin receptor‐mediated signal transduction in rat stomach

FEBS Letters

Takii, Takemasa¹  Ohya, Susumu²  Matsumori, Miki²  Onozaki, Kikuo¹  Yamazaki, Hiroko F.²  Imaizumi, Yuji²  Watanabe, Minoru² 

[1] Department of Hygienic Chemistry, Faculty of Pharmaceutical Sciences, Nagoya City University, 3-1 Tanabedori, Mizuhoku, Nagoya 467, Japan;Department of Chemical Pharmacology, Faculty of Pharmaceutical Sciences, Nagoya City University, 3-1 Tanabedori, Mizuhoku, Nagoya 467, Japan

关键词: 5-HT receptor;    cDNA cloning;    Xenopus oocyte;    In vitro translation;    Northern blotting;    [3H]Spiperone binding assay;   

DOI  :  10.1016/S0014-5793(96)01485-8

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

In Xenopus oocytes injected with small size mRNAs (500–700 b), obtained from rat stomach by fractionation, application of 10 μM 5-HT induced a substantial Ca²⁺-activated Cl⁻ current (I_Cl-Ca). I_Cl-Ca was not elicited by 5-HT in native oocytes. Consistent results from this assay in the oocyte expression system motivated cDNA cloning experiments. A novel cDNA (namedattomacherotonin receptor-related cDNA: RSS cDNA) which encodes a small protein involved in specific 5-HT receptor-mediated I_Cl-Ca activation was identified. The molecular weight of RSS protein in the reticulocyte lysate translation system (∼10 kDa) is identical to that calculated from the amino acid sequence. Computer-aided analysis of the predicted protein does not show any obvious sequence homologies (<18%) to any other proteins including G protein-coupled receptors. Northern analysis revealed that RSS mRNA is ubiquitously expressed at varying levels in a number of different tissues. Furthermore, the binding of [³H]spiperone, a 5-HT₂ receptor antagonist, was examined in CHO cells, which highly expressed RSS transcripts (named CHO-RSS). Specific binding of [³H]spiperone was not clearly observed in native CHO but was detected in CHO-RSS. The dissociation constant was 10.3 nM in CHO-RSS. These results suggest that RSS protein may be a factor which facilitates 5-HT receptor expression or, alternatively, an enhancer of the affinity of native 5-HT receptor to 5-HT.

【 授权许可】

Unknown