Dynamical Structural Change of Reconstituted Chromatin and Its Genetic Activity

重组染色质的动态结构变化及其遗传活性

基本信息

批准号：
14598005
负责人：
YOSHIKAWA Yuko
金额：
$ 2.24万
依托单位：
College of Nagoya Bunri University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2004
项目状态：
已结题

项目摘要

Recently, the methods of single-molecule observation have been utilized to study molecular mechanisms of biological reactions in vitro. With fluorescence microscopy, I have made direct observations of single giant DNA molecules in solution and have found that giant DNA molecules change their conformation depending on environmental conditions.The main results in the present study are summarized as follows.(1) I studied the effect of daunomycin, a cancer chemotherapeutic agent, on a model cellular system in which folded compact DNA was encapsulated inside a cell-sized phospholipid liposome. Following the exposure to daunomycin, the compact DNA inside a liposome was gradually elongated and then cut into a pair of fragments. It has become clear that daunomycin can penetrate the bilayer lipid membrane and has the potential effect to loosen the packing of DNA together with the activity to cut the double stranded DNA. Our results imply that cell-sized liposomes are useful as a simple model of living cells.(2) I examined the effect of ascorbic acid on the higher order structure of DNA through single molecular observation with fluorescence microscopy, and found that ascorbic acid generates a pearling structure in giant DNA molecules, where elongated and compact parts coexist along a molecular chain. The results of observations with atomic force microscopy indicate that the compact parts assume a loosely packed conformation. It has been reported that human circulating immune cells, such as neutrophils, monocytes and lymphocytes, accumulate ascorbic acid in millimolar concentrations. Therefore, it is expected that the ascorbic acid concentration that induces the large conformational change on DNA may be of physiological significance.(3) I investigated the effect of ascorbic acid on preventing DNA double-strand breaks in reconstituted chromatin in relation to the highly compacted polynucleosomal structure.

近年来，单分子观察的方法已被用来研究体外生物反应的分子机制。通过荧光显微镜，我对溶液中的单个巨型DNA分子进行了直接观察，发现巨型DNA分子会根据环境条件改变其构象。本研究的主要结果总结如下：（1）我研究了这种效应道诺霉素（一种癌症化疗剂）在模型细胞系统中的应用，其中折叠的紧凑 DNA 被封装在细胞大小的磷脂脂质体内。接触道诺霉素后，脂质体内的紧凑 DNA 逐渐拉长，然后切割成一对片段。现已明确，道诺霉素可以穿透双层脂膜，具有松散DNA堆积的潜在作用以及切割双链DNA的活性。我们的结果表明细胞大小的脂质体可用作活细胞的简单模型。(2) 我通过荧光显微镜的单分子观察检查了抗坏血酸对 DNA 高级结构的影响，发现抗坏血酸产生巨型 DNA 分子中的珍珠结构，其中细长部分和紧凑部分沿着分子链共存。原子力显微镜的观察结果表明，致密部分呈现松散堆积的构象。据报道，人类循环免疫细胞，如中性粒细胞、单核细胞和淋巴细胞，会积聚毫摩尔浓度的抗坏血酸。因此，预计诱导DNA大构象变化的抗坏血酸浓度可能具有生理意义。(3)我研究了抗坏血酸在防止重构染色质中DNA双链断裂的作用，这与高度压缩的染色质有关。多核小体结构。

项目成果

期刊论文数量（43）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（0）

All-or-none folding transition in giant mammalian DNA

巨型哺乳动物DNA中的全或无折叠转变

DOI：
发表时间：
2002
期刊：
Chem.Phys.Lett. 354
影响因子：
0
作者：
Kenichi Yoshikawa
通讯作者：
Kenichi Yoshikawa

Daunomycin triggers membrane blebbing and breakage of giant DNA encapsulated in a cell-sized liposome

道诺霉素引发细胞大小脂质体中包裹的巨型 DNA 的膜起泡和断裂

DOI：
发表时间：
2002
期刊：
Chem.Phys.Lett. 366
影响因子：
0
作者：
Yuko Yoshikawa
通讯作者：
Yuko Yoshikawa

巨大DNA分子の折り畳み転移:高次構造と機能

DNA 大分子的折叠转变：高阶结构和功能

DOI：
发表时间：
2003
期刊：
放射線生物研究 38
影响因子：
0
作者：
Noriko Takenaka;Yuko Yoshikawa;Kumiko Hibino;Y.Yoshikawa et al.;K.Hibino et al.;Kumiko Hibino;Yuko Yoshikawa;吉川祐子
通讯作者：
吉川祐子

Folding Transition of Large DNA Completely Inhibits the Action of a Restriction Endonuclease as Revealed by Single-Chain Observation

单链观察表明，大 DNA 的折叠转变完全抑制限制性内切酶的作用