Scuric Zorica, Chan Cecilia Y, Hafer Kurt, Schiestl Robert H
David Geffen School of Medicine at UCLA, Department of Pathology, Los Angeles, California, USA.
Radiat Res. 2009 Apr;171(4):454-63. doi: 10.1667/RR1329.1.
DNA double-strand breaks repaired through nonhomologous end joining require no extended sequence homology as a template for the repair. A subset of end-joining events, termed microhomology-mediated end joining, occur between a few base pairs of homology, and such pathways have been implicated in different human cancers and genetic diseases. Here we investigated the effect of exposure of yeast and mammalian cells to ionizing radiation on the frequency and mechanism of rejoining of transfected unirradiated linear plasmid DNA. Cells were exposed to gamma radiation prior to plasmid transfection; subsequently the rejoined plasmids were recovered and the junction sequences were analyzed. In irradiated yeast cells, 68% of recovered plasmids contained microhomologies, compared to only 30% from unirradiated cells. Among them 57% of events used>or=4 bp of microhomology compared to only 11% from unirradiated cells. In irradiated mammalian cells, 54% of plasmids used>or=4 bp of microhomology compared to none from unirradiated cells. We conclude that exposure of yeast and mammalian cells to radiation prior to plasmid transfection enhances the frequency of microhomology-mediated end-joining events in trans. If such events occur within genomic locations, they may be involved in the generation of large deletions and other chromosomal aberrations that occur in cancer cells.
通过非同源末端连接修复的DNA双链断裂不需要延伸的序列同源性作为修复模板。一部分末端连接事件，称为微同源性介导的末端连接，发生在少数几个碱基对的同源序列之间，并且这种途径与不同的人类癌症和遗传疾病有关。在这里，我们研究了酵母细胞和哺乳动物细胞暴露于电离辐射对转染的未受辐射的线性质粒DNA重新连接频率和机制的影响。在质粒转染之前，细胞先暴露于γ辐射；随后回收重新连接的质粒并分析连接序列。在受辐射的酵母细胞中，68%回收的质粒含有微同源序列，而未受辐射的细胞中这一比例仅为30%。其中，57%的事件使用了≥4个碱基对的微同源序列，而未受辐射的细胞中这一比例仅为11%。在受辐射的哺乳动物细胞中，54%的质粒使用了≥4个碱基对的微同源序列，而未受辐射的细胞中未出现这种情况。我们得出结论，在质粒转染之前，酵母细胞和哺乳动物细胞暴露于辐射会提高微同源性介导的反式末端连接事件的频率。如果这些事件发生在基因组位置内，它们可能参与癌细胞中出现的大缺失和其他染色体畸变的产生。
