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Roles of vasoactive intestinal contractor/endothelin-2 in bovine ovary

血管活性肠收缩剂/内皮素-2 在牛卵巢中的作用

基本信息

批准号：
13660308
负责人：
UCHIDE Tsuyoshi
金额：
$ 2.18万
依托单位：
Kitasato University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2003
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13660308/
关键词：
endothelin vasoactive intestinal contractor bovine ovary cDNA cloning mRNA expression real-time PCR organ distribution of mRNA polypeptide sequence ウシエンドセリン2 クローニング卵巣

项目摘要

Endothelia-2(ET2), which was originally identified in human, is a bioactive peptide of 21 amino acids with strong vasoconstrictive and pressor effects. Here we report the cDNA cloning and characterization of bovine preproendothelin-2(PPET2), the precursor form of ET2. The bovine cDNA encodes 177 amino acids of the PPET2 polypeptide, in which a 21-amino acid mature ET2 peptide and a 16-amino acid ET2-like peptide as well as a 23-amino acid putative signal peptide were found. The bovine ET2-like peptide sequence was missing a dibasic amino acid pair at the C-terminal, in contrast to human, mouse and rat, for which the ET2-like sequence flanked dibasic pairs at both the N-and C-terminals. Gene expression analysis by RT-PCR showed that the transcript is expressed in various organs including heart, lung, liver, kidney, gastrointestinal tract, uterus and ovary, but not in spleen. Within the gastrointestinal tract, gene expression was detected in rumen, a ruminant-specific digestive organ, as well as stomach, duodenum and colon. Based on the full-length bovine PPET2 cDNA sequence obtained, we designed gene-specific primers and TagMan probe, and then established a rapid and precise quantitative method for the gene expression using a real-time PCR system.

内皮素-2（ET-2）是一种由21个氨基酸组成的生物活性肽，具有强的缩血管和升压作用。在这里，我们报告的cDNA克隆和表征牛前原内皮素-2（PPET 2），ET 2的前体形式。牛cDNA编码PPET 2多肽的177个氨基酸，其中发现了一个21个氨基酸的成熟ET 2肽和一个16个氨基酸的ET 2样肽以及一个23个氨基酸的推定信号肽。牛ET 2样肽序列在C端缺失一个二元氨基酸对，与人、小鼠和大鼠相反，其中ET 2样序列在N-和C-末端都侧接二碱基对。RT-PCR分析表明，该基因在心、肺、肝、肾、胃肠道、子宫和卵巢等器官中均有表达，但在脾脏中不表达。在胃肠道内，基因表达在瘤胃，反刍动物特有的消化器官，以及胃，十二指肠和结肠中检测到。根据获得的牛PPET 2全长cDNA序列，设计了基因特异性引物和TagMan探针，建立了一种快速、准确的实时荧光定量PCR方法。