Liu Z, Hall J D, Mount D W
Department of Molecular and Cellular Biology, University of Arizona, Tucson, Arizona 85721, USA.
Plant J. 2001 May;26(3):329-38. doi: 10.1046/j.1365-313x.2001.01031.x.
To identify mechanisms of DNA repair in Arabidopsis thaliana, we have analyzed a mutant (uvh3) which exhibits increased sensitivity to ultraviolet (UV) light, H2O2 and ionizing radiation and displays a premature senescence phenotype. The uvh3 locus was mapped within chromosome III to the GL1 locus. A cosmid contig of the GL1 region was constructed, and individual cosmids were used to transform uvh3 mutant plants. Cosmid N9 was found to confer UV-resistance, H2O2-resistance and a normal senescence phenotype following transformation, indicating that the UVH3 gene is located on this cosmid and that all three phenotypes are due to the same mutation. Analysis of cosmid N9 sequences identified a gene showing strong similarity to two homologous repair genes, RAD2 (Saccharomyces cerevisiae) and XPG (human), which encode an endonuclease required for nucleotide excision repair of UV-damage. The uvh3 mutant was shown to carry a nonsense mutation in the coding region of the AtRAD2/XPG gene, thus revealing that the UVH3 gene encodes the AtRAD2/XPG gene product. In humans, the homologous XPG protein is also involved in removal of oxygen-damaged nucleotides by base excision repair. We discuss the possibility that the increased sensitivity of the uvh3 mutant to H2O2 and the premature senescence phenotype might result from failure to repair oxygen damage in plant tissues. Finally, we show that the AtRAD2/XPG gene is expressed at moderate levels in all plant tissues.
为了确定拟南芥中DNA修复的机制,我们分析了一个突变体(uvh3),该突变体对紫外线(UV)、H2O2和电离辐射表现出更高的敏感性,并呈现出早衰表型。uvh3基因座被定位在第三条染色体上的GL1基因座内。构建了GL1区域的黏粒重叠群,并使用单个黏粒转化uvh3突变体植株。发现黏粒N9在转化后赋予了抗紫外线、抗H2O2能力和正常的衰老表型,这表明UVH3基因位于该黏粒上,并且所有这三种表型都是由同一个突变引起的。对黏粒N9序列的分析鉴定出一个与两个同源修复基因RAD2(酿酒酵母)和XPG(人类)具有高度相似性的基因,这两个基因编码紫外线损伤核苷酸切除修复所需的一种核酸内切酶。uvh3突变体被证明在AtRAD2/XPG基因的编码区携带一个无义突变,从而揭示UVH3基因编码AtRAD2/XPG基因产物。在人类中,同源的XPG蛋白也参与碱基切除修复去除氧化损伤的核苷酸。我们讨论了uvh3突变体对H2O2敏感性增加和早衰表型可能是由于植物组织中氧化损伤修复失败导致的可能性。最后,我们表明AtRAD2/XPG基因在所有植物组织中均以中等水平表达。