Spectra of X-ray-induced and spontaneous intragenic HPRT mutations in closely related human cells differentially expressing the p53 tumor suppressor gene.
Spectra of X-ray-induced and spontaneous intragenic HPRT mutations in closely related human cells differentially expressing the p53 tumor suppressor gene.
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DOI：
10.2307/3579414
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发表时间：
1997-02
影响因子：
3.4
通讯作者：
E. Phillips;D. Gebow;H. Liber
中科院分区：
文献类型：
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作者：
E. Phillips;D. Gebow;H. Liber
先前对两种密切相关的人淋巴母细胞系的研究表明，WTK1细胞比TK6细胞对X射线诱导的细胞毒性更具抗性，但在TK和HPRT基因座上更具突变性。随后确定WTK 1细胞过表达肿瘤抑制基因p53的突变形式，而TK 6细胞正确表达野生型p53。因此，这两个细胞系使我们能够检查在HPRT基因座的突变谱在相关的人类淋巴母细胞细胞系差异表达p53。在此之前，我们分离了X射线诱导的和自发的WTK 1突变体和自发的TK 6突变体，并通过多重聚合酶链反应（PCR）和Southern印迹分析相结合的突变谱进行了分析。有些出乎意料的是，我们发现，尽管X射线在WTK 1细胞中诱导的突变体大约是TK 6细胞中的四倍，但突变谱几乎没有差异。在本研究中，为了确定X射线诱导的WTK 1突变体中是否存在更高频率的基因内缺失，我们进一步检查了含有HPRT点突变的突变体子集。cDNA序列分析被用来精确地确定在19个X射线诱导的和25个自发的WTK 1突变体和25个自发的TK 6突变体的突变。虽然这两种细胞系之间的HPRT突变谱存在细微差异，但数据再次表明，p53与HPRT突变频率的增加相关，而对恢复的突变类型没有明显影响。
Previous work with two closely related human lymphoblast cell lines demonstrated that WTK1 cells are more resistant than TK6 cells to X-ray-induced cytotoxicity, but more mutable at both the TK and HPRT loci. It was subsequently determined that WTK1 cells overexpress a mutant form of the tumor suppressor gene p53, while TK6 cells correctly express wild-type p53. Thus these two cell lines allowed us to examine the mutational spectra at the HPRT locus in related human lymphoblast cell lines differentially expressing p53. Previously, we isolated sets of X-ray-induced and spontaneous WTK1 mutants and spontaneous TK6 mutants and analyzed the mutational spectra by the combination of multiplex polymerase chain reaction (PCR) and Southern blot analysis. Somewhat unexpectedly, we found that even though there were approximately four times as many mutants induced by X rays in WTK1 cells as in TK6 cells, there was very little difference in the mutational spectra. In the present study, to determine if there was a higher frequency of intragenic deletions among the X-ray-induced WTK1 mutants, we further examined the subsets of mutants that contained HPRT point mutations. cDNA sequence analysis was used to define the mutation precisely in 19 X-ray-induced and 25 spontaneous WTK1 mutants and 25 spontaneous TK6 mutants. While subtle differences exist in the spectra of HPRT mutations between these two cell lines, the data again suggest that p53 is associated with an increase in the frequency of mutations at HPRT without an obvious effect on the types of mutations recovered.