Cloning of apoptosis-inducing genes by introducing genes into living tadpole

通过将基因导入活体蝌蚪来克隆凋亡诱导基因

基本信息

批准号：
10670129
负责人：
YAOITA Yoshio
金额：
$ 1.92万
依托单位：
Hiroshima University (2000)Tokyo Metropolitan Organization for Medical Research (1999)Tokyo Metropolitan Institute for Neuroscience (1998)
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 2000
项目状态：
已结题

项目摘要

Our purpose is to clone apoptosis-inducing genes that are induced by thyroid hormone (TH) in the regressing tail of tadpole during amphibian metamorphosis.1. We have isolated all Xenopus Caspase family genes as apoptosis-excuting genes, and introduced them into tail-derived myoblast cell line. The overexpression of Caspases with a long prodomain showed a strong cell-killing activity. The expression of Xenopus Caspase family genes was upregulated in the regressing tail of metamorphosing tadpole, but was not in dying tail-derived cell line treated with TH, which demonstrates that new RNA synthesis of caspases is dispensable to programmed cell death.2. Many reports supported the murder model that the increase of secreted extracellular matrixdegrading enzymes induced by TH results in the disruption of the myotendinous junctions, which detaches muscle cells from the extracellular matrix and causes their cell death. However, when TH-signaling was blocked by the enforced expression of dominan … More t-negative thyroid hormone receptor (DNTR), the death of TH-treated myoblast cell line was repressed, suggesting that the programmed cell death takes place through a suicide mechanism. To confirm it, DNTR expression construct was injected into the tail muscle cells of living tadpole with a marker gene, and the activity of marker protein was detected just before tail resorbed completely. The marker gene expression in muscle cells was observed only in tails co-injected with DNTR expression construct. This result indicates that muscle cell dies by suicide as a response to TH in the regressing tail.3. Tadpole tail was introduced with a marker gene and cDNA library in a expression vector using in vivo electroporation, cut out for the organ culture several days later, and examined on the maker gene expression. Any clone was not isolated as one inhibiting marker gene expression. The overexpression of several genes identified so far by the increased expression in the regressing tail did not show any suppressive activity, either. We are analyzing a group of cDNA clones which has the significant suppressive activity to the marker gene expression in the screening by transfection of tail-derived mvoblastic cell line. Less

我们的甲状腺激素（Th）在两栖动物的t尾中诱导甲状腺激素（Th），我们将所有Xenopus caspase家族作为凋亡型基因，并将其引入尾巴衍生的肌色细胞系用长prodomain的Caspass过表达表现出强大的细胞杀伤活性。 .2。通过自杀机制进行编程细胞，以确认与标记基因的t，并且在尾巴压缩的尾巴压缩之前检测到标记蛋白的活性。在回归尾巴中，SE使用体内电穿孔的表达载体，几天后，降低了器官培养的严重程度，而不是作为抑制标记基因表达的一种。我们没有显示任何抑制活性