生物谷： 復(fù)制誘導(dǎo)的DNA斷裂是一個自發(fā)過程，有可能是染色體重組引起癌癥的主要起因,。但是,，在正常細(xì)胞中，這個過程的發(fā)生概率有多大還不清楚,。一項新的研究表明,，這個過程發(fā)生的概率非常低。然而,，一旦發(fā)生,，它對基因組穩(wěn)定性的危害將是之前預(yù)計的100倍。
在大腸桿菌的環(huán)形基因組上,，攜帶著復(fù)制起點oriC（左上圖）。一旦復(fù)制啟動（右上圖）,，將形成兩個復(fù)制叉（桔色的三角形）,。其中的一個復(fù)制叉停滯了下來（右下圖），并且發(fā)生瓦解（復(fù)制蛋白解體,，從DNA上脫落下來,，圖中用空的三角形表示）。在瓦解的復(fù)制叉上會產(chǎn)生一個DSB,，并由之誘導(dǎo)SOS DNA損傷反應(yīng),。這個反應(yīng)誘導(dǎo)轉(zhuǎn)錄從sulA開始（桔色的四邊形），誘導(dǎo)gfp基因的表達(dá),。在誘導(dǎo)SOS反應(yīng)后,，大腸桿菌呈現(xiàn)綠色（左下圖）。此時,，細(xì)胞可以修復(fù)瓦解的復(fù)制叉,，并繼續(xù)進(jìn)行復(fù)制。
原始出處:
Nature Genetics 39, 797 - 802 (2007)
Published online: 27 May 2007 | doi:10.1038/ng2051
Spontaneous DNA breakage in single living Escherichia coli cells
Jeanine M Pennington1 & Susan M Rosenberg1,2
Abstract
Spontaneous DNA breakage is predicted to be a frequent, inevitable consequence of DNA replication and is thought to underlie much of the genomic change that fuels cancer and evolution1, 2, 3. Despite its importance, there has been little direct measurement of the amounts, types, sources and fates of spontaneous DNA lesions in living cells. We present a direct, sensitive flow cytometric assay in single living Escherichia coli cells for DNA lesions capable of inducing the SOS DNA damage response, and we report its use in quantification of spontaneous DNA double-strand breaks (DSBs). We report efficient detection of single chromosomal DSBs and rates of spontaneous breakage 20- to 100-fold lower than predicted. In addition, we implicate DNA replication in the origin of spontaneous DSBs with the finding of fewer spontaneous DSBs in a mutant with altered DNA polymerase III. The data imply that spontaneous DSBs induce genomic changes and instability 20–100 times more potently than previously appreciated. Finally, FACS demonstrated two main cell fates after spontaneous DNA damage: viability with or without resumption of proliferation.
Interdepartmental Program in Cell and Molecular Biology and Department of Molecular and Human Genetics, Houston, Texas 77030-3411, USA.
Department of Biochemistry and Molecular Biology, Department of Molecular Virology and Microbiology and Dan L. Duncan Cancer Center, Baylor College of Medicine, One Baylor Plaza, Houston, Texas 77030-3411, USA.
Correspondence to: Susan M Rosenberg1,2 e-mail: [email protected]
