Ziv Y, Bar-Shira A, Jorgensen T J, Russell P S, Sartiel A, Shows T B, Eddy R L, Buchwald M, Legerski R, Schimke R T, Shiloh Y
Department of Human Genetics, Sackler School of Medicine, Tel Aviv University, Ramat Aviv, Israel.
Somat Cell Mol Genet. 1995 Mar;21(2):99-111. doi: 10.1007/BF02255785.
Genes responsible for genetic diseases with increased sensitivity to DNA-damaging agents can be identified using complementation cloning. This strategy is based on in vitro complementation of the cellular sensitivity by gene transfer. Ataxia-telangiectasia (A-T) is a multisystem autosomal recessive disorder involving cellular sensitivity to ionizing radiation and radiomimetic drugs. A-T is genetically heterogeneous, with four complementation groups. We attempted to identify cDNA clones that modify the radiomimetic sensitivity of A-T cells assigned to complementation group [A-T(A)]. The cells were transfected with human cDNA libraries cloned in episomal vectors, and various protocols of radiomimetic selection were applied. Thirteen cDNAs rescued from survivor cells were found to confer various degrees of radiomimetic resistance to A-T(A) cells upon repeated introduction, and one of them also partially influenced another feature of the A-T phenotype, radioresistant DNA synthesis. None of the clones mapped to the A-T locus on chromosome 11q22-23. Nine of the clones were derived from known genes, some of which are involved in cellular stress responses. We concluded that a number of different genes, not necessarily associated with A-T, can influence the response of A-T cells to radiomimetic drugs, and hence the complementation cloning approach may be less applicable to A-T than to other diseases involving abnormal processing of DNA damage.
利用互补克隆可鉴定出对DNA损伤剂敏感性增加的遗传性疾病相关基因。该策略基于通过基因转移对细胞敏感性进行体外互补。共济失调毛细血管扩张症(A-T)是一种多系统常染色体隐性疾病,涉及细胞对电离辐射和放射模拟药物的敏感性。A-T在遗传上具有异质性,有四个互补组。我们试图鉴定可改变分配到互补组[A-T(A)]的A-T细胞对放射模拟物敏感性的cDNA克隆。用克隆于附加型载体的人cDNA文库转染细胞,并应用各种放射模拟物选择方案。从存活细胞中挽救出的13个cDNA经反复导入后,发现可赋予A-T(A)细胞不同程度的放射模拟物抗性,其中一个还部分影响了A-T表型的另一个特征,即抗辐射DNA合成。这些克隆均未定位于11号染色体q22-23上的A-T基因座。其中9个克隆来自已知基因,有些基因参与细胞应激反应。我们得出结论,许多不同的基因(不一定与A-T相关)可影响A-T细胞对放射模拟药物的反应,因此互补克隆方法对A-T的适用性可能低于对其他涉及DNA损伤异常处理的疾病。