Molecular Biology of the Biological Clock : From Gene to Behavior

生物钟的分子生物学：从基因到行为

基本信息

批准号：
11470018
负责人：
OKAMURA Hitoshi
金额：
$ 9.66万
依托单位：
Kobe University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B).
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2000
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-11470018/
关键词：
mPer1 clock genes dbp cry e4bp4 circadian rhythm transgenic mice suprachiasmatic nucleus 哺乳類時計遺伝子サーカディアンリズム Cry 単一細胞のリズム

项目摘要

In the eukaryotic circadian model systems, translocation of the oscillatory gene products into the nucleus is a key step for generation of a 24 hour cycle of the biological clock. We have examined nuclear import of clock proteins of the mammalian period gene family and the effect of serum shock, which induces a synchronous clock in cultured cells. We examined the nuclear import of mPER proteins in COS7 cells and found that nuclear translocation of mPER1 and mPER2 involves physical interactions with mPER3. This indicates that nuclear translocation of mPER1 also can occur under physiological conditions in the absence of CRY proteins.Recently we demonstrated that an accessory transcription loop exists helping to the core clock feedback loop. Transcript levels of DBP, a member of the PAR leucine zipper transcription factor family, exhibit a robust rhythm in the SCN.We report that DBP is able to activate the promoter of a putative clock oscillating gene, mPer1, by directly binding to the mPer1 promoter. DBP and CLOCK-BMAL1 cooperatively activate the mPer1 promoter. On the other hand, dbp transcription is activated by CLOCK-BMAL1 through E-boxes and inhibited by the mPER and mCRY proteins, as is the case for mPer1. Antiphase circadian regulated E4BP4, another member of leucine zipper transcription factor without PAR domain, antagonistically suppresses mPer1 transcription. Thus, a clock-controlled dbp and e4bp4 genes may play an important role in the central clock oscillation.Using transgenic mice carrying the mPer1 promoter fused to the luciferase (mPer1-luc) gene, we recently succeeded to monitor luciferase-mediated bioluminescence with a day-night variation in the SCN in brain slices and in living animals. We can record for several days oscillating photon emission with a periodicity and phase that accurately mirrored native mPer1 mRNA expression. The real-time optical imaging of gene expression will be a new powerful tool to study mammalian brain function.

在真核生物昼夜节律模型系统中，振荡基因产物转移到核中是生成生物钟24小时周期的关键步骤。我们已经检查了哺乳动物周期基因家族的核进口核进口蛋白以及血清休克的作用，血清休克的作用诱导了培养细胞中的同步时钟。我们检查了MPER蛋白在COS7细胞中的核进口，发现MPER1和MPER2的核转运涉及与MPER3的物理相互作用。这表明在没有哭泣蛋白的情况下，MPER1的核易位也可以在生理条件下发生。我们证明，存在辅助转录环有助于核心时钟反馈回路。 DBP的转录水平是PAR亮氨酸拉链转录因子家族的成员，在SCN中表现出强大的节奏。我们报告说，DBP能够通过直接与MPER1启动子结合来激活推定的时钟振荡基因MPER1的启动子。 DBP和Clock-BMAL1合作激活MPER1启动子。另一方面，DBP转录被Clock-BMAL1通过电子盒激活，并被MPER和MCRY蛋白抑制，MPER1也是如此。反相昼夜节律调节的E4BP4，这是没有par结构域的亮氨酸拉链转录因子的另一个成员，可以拮抗MPER1转录。因此，时钟控制的DBP和E4BP4基因可能在中央时钟振荡中起重要作用。使用携带MPER1启动子与荧光素酶（MPER1-LUC）基因的转基因小鼠，我们最近成功地监测了Luciferase介导的生物发光，并在动物中监测了日夜在SCN中的日夜变异。我们可以通过周期性和相位准确地反映了天然MPER1 mRNA表达的周期性和相位来记录几天的振荡光子发射。基因表达的实时光学成像将是研究哺乳动物脑功能的新型工具。

项目成果

期刊论文数量（76）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

Takumi T, Nagamine Y, Miyake S, Matsubara C, Taguchi K, Takekida S, Sakakida Y, Nishikawa K, Kishimoto T, Niwa S, Okumura K, Okamura H: "A mammalian orthologue of Drosophila timeless, highly expressed in SCN and retina, forms a complex with mPER1"Genes Ce

Takumi T、Nagamine Y、Miyake S、Matsubara C、Taguchi K、Takekida S、Sakakida Y、Nishikawa K、Kishimoto T、Niwa S、Okumura K、Okamura H：“果蝇永恒的哺乳动物直系同源物，在 SCN 和视网膜中高度表达