Identification of Oxygen Sensor Expressed in Newborn Rats

新生大鼠中表达的氧传感器的鉴定

• 批准号: 10671025
• 负责人: TAKAHASHI Yuji
• 金额: $ 1.98万
• 依托单位: Tokyo University of Pharmacy and Life Science
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10671025/
• 关键词: hypoxia; oxygen sensor; differential display; fibroblast; prolyl 4-hydroxylase; rat; fetus; 酸素; センサー; エリスロポイエチン; GFP; Ba; F3; 新生児; 呼吸器; 循環器

Molecular oxygen (O2) is essential for aerobic organisms. Exposure of tissues or cells to hypoxia induces a variety of adaptive or pathogenic responses. Accumulated evidence indicates that hypoxia activities collagen synthesis in tissues. To explore the molecular mechanism we screened those genes which are up-regulated or down-regulated by hypoxia. Fibroblasts isolated from fetal rat lung were cultured under hypoxia. Differential display technique showed that the mRNA level of prolyl 4-hydroxylase (PH) a(I), an active subunit which catalyzes the oxygen-dependent hydroxylation of proline residue in procollagen, was increased by 2 to 3 fold after an 8 h exposure to hypoxia. This elevated level was maintained over 40 h, and returned to the basal level after reoxygenation. The transcription rate, the protein level and the hydroxy proline content, an indicator of the prolyl hydroxylation, were all elevated by hypoxic culture. Analysis of the promotor region of PHa(I) gene indicated that a motif similar to hypoxia responsive element (HRE) of hypoxia-inducible genes such as erythropoietin was identified within 120-base pair sequence upstream the transcription start site. Luciferase-reporter assay and mutational analysis showed that a site similar to the HRE in this motif is functionally essential to hypoxic response. Electrophoretic mobility shift assay revealed that hypoxia inducible factor-1 (HIF-1) was stimulated and bound to the PHa(I) HRE upon hypoxic challenge. Our results indicate that PHa(I), an essential enzyme for collagen synthesis, is a target gene for HIF-1.

分子氧（O2）是好氧生物所必需的。组织或细胞暴露于缺氧诱导多种适应性或致病性反应。越来越多的证据表明，缺氧促进组织中胶原蛋白的合成。为了探讨缺氧的分子机制，我们筛选了缺氧上调或下调的基因。从胎鼠肺分离的成纤维细胞在缺氧条件下培养。差异显示技术显示，缺氧8h后，脯氨酰4-羟化酶（PH）a（I）的mRNA水平增加2 ~ 3倍。这种升高的水平维持了40小时以上，并在复氧后恢复到基础水平。低氧培养提高了转录速率、蛋白质水平和脯氨酰羟化的指标羟脯氨酸含量。对PHa（I）基因启动子区的分析表明，在转录起始位点上游120个碱基对的序列中发现了一个与促红细胞生成素等低氧诱导基因的低氧反应元件（HRE）相似的基序。荧光素酶报告分析和突变分析表明，在这个基序的HRE类似的网站是功能上必不可少的缺氧反应。电泳迁移率改变实验显示缺氧诱导因子-1（HIF-1）在缺氧刺激下被激活并与PHa（I）HRE结合。我们的研究结果表明，PHa（I），胶原合成的必需酶，是HIF-1的靶基因。

项目成果

Takahashi, Y, et al: "Oxygen tension regulates heme oxygenase-1"Cell Biochem. Function. 16. 183-193 (1998)

Takahashi, Y 等人：“氧张力调节血红素加氧酶-1”Cell Biochem。

Takahashi. S. et al: "Cell type-depenclent regulatory elements."Biochim. Biophy. Actce. 1447. 231-235 (1999)

高桥。

Y.Takahashi ら: "Mypox a indrced expression of phosphogyierate mutnso B in fibroblasts" Eur. J. Biochem.254. 497-504 (1998)

Y. Takahashi 等：“Mypox 诱导成纤维细胞中磷酸甘油酸 mutnso B 的表达”Eur. J. Biochem.254 (1998)。

Takahashi, Y, et al: "Hypoxic induction of prolyl 4-hydroxylase"J. Biol. chem. (in press). (2000)

Takahashi, Y, et al：“脯氨酰 4-羟化酶的缺氧诱导”J。

Yuji Takahashi, Shigeru Takahashi, Tatsuya Yoshimi, Takashi Miura: "Hypoxia induced expression of phosphoglycerate mutase B in fibroblasts."Eur.J.Biochem.. 254. 497-504 (1998)

Yuji Takahashi、Shigeru Takahashi、Tatsuya Yoshimi、Takashi Miura：“缺氧诱导成纤维细胞中磷酸甘油酸变位酶 B 的表达。”Eur.J.Biochem.. 254. 497-504 (1998)