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Involvement of p73-dependent apoptosis pathway in the process of acquisition of drug-resistance in human lung cancer cell lines

p73依赖性凋亡途径参与人肺癌细胞系耐药获得过程

基本信息

批准号：
12470133
负责人：
HAGIWARA Koichi
金额：
$ 8.77万
依托单位：
Tohoku University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2001
项目状态：
已结题

项目摘要

p73 is a candidate tumor suppressor gene with substantial DNA and protein homology to the p53 tumor suppressor gene. In the first part of this study, we have investigated two hypotheses: (a) p73 is mutated in diverse types of human cancer, and (b) p73 is functionally redundant with p53 in carcinogenesis so that mutations would be exclusive in these two genes. The entire coding region and intronic splice junctions of p73 were examined in 54 cancer cell lines. Three lung cancer cell lines contained mutations that affected the amino acid sequence. One amino acid substitution was in a region with homology to the specific DNA binding region of p53 and two microdeletions were outside the region of homology. Two of the cell lines with p73 mutations also carried p53 mutations. Our results are inconsistent with the two hypotheses tested. Next we checked hypotheses for p63 that (a) p63 is mutated in diverse types of human cancers and (b)p63 functions in the same pathway as p53 and p73 in the pro … More cess of carcinogenesis, so that mutations in these three genes would be mutually exclusive. We analyzed the genomic structure of the p63 gene and have performed mutational analyses using the same set of cell lines as for p73. We have shown that DLD1 and SKOV3 cells have either heterozygous mutations or polymorphisms in the putative DNA binding domain of p63. In these cell lines, p63 is biallelically expressed. In the last part of the study, we analyzed three naturally occurring p73 mutants found in lung cancer cell lines. NCI-H1155 has a p73 mutation, p73(G264W), in the DNA binding domain, as well as a "gain-of-function" p53 mutation, p53(R273H). p73α(G264W) lacks the transactivation activity itself, and suppressed the transactivation activity of wild-type p73α, indicating that p73α(G264W) is a dominant negative mutant. Consistently, p73α(G264W) failed to suppress colony formation. p73 mutants found in DMS 92 or in A427 showed no functional abnormalities. In NCI-H1155 cells the coexistence of mutations that abrogate the normal function of p73 and p53 may indicate that each mutation confers an additive growth advantage on the cells. Less

P73是一个候选抑癌基因，与P53抑癌基因在DNA和蛋白质上具有很高的同源性。在这项研究的第一部分，我们研究了两个假设：(A)p73在不同类型的人类癌症中发生突变，以及(B)p73在致癌过程中与p53在功能上是多余的，因此这两个基因中的突变是唯一的。在54个癌细胞系中检测了p73的整个编码区和内含子剪接连接。三个肺癌细胞系含有影响氨基酸序列的突变。1个氨基酸替换位于与P53特异DNA结合区同源的区域，2个微缺失位于同源区域之外。其中两个带有p73突变的细胞系也带有p53突变。我们的结果与测试的两个假设不一致。接下来，我们验证了p63的假设，即(A)p63在不同类型的人类癌症中发生突变，以及(B)p63在PRO…中的功能与p53和p73相同致癌的几率更大，因此这三个基因的突变是相互排斥的。我们分析了p63基因的基因组结构，并使用与p73相同的细胞系进行了突变分析。我们已经证明，DLD1和SKOV3细胞在p63的DNA结合域存在杂合突变或多态。在这些细胞系中，p63是双等位表达的。在研究的最后部分，我们分析了在肺癌细胞系中发现的三个自然发生的p73突变。NCI-H1155在DNA结合区有p73突变p73(G264W)，以及“功能获得”的p53突变P53(R273H)。P73α(G264W)本身缺乏反式激活活性，并抑制野生型p73α的反式激活活性，表明p73α(G264W)为显性负性突变体。P73α(G264W)不能抑制克隆形成。在DMS 92和A427中发现的p73突变没有表现出功能异常。在NCI-H1155细胞中，破坏p73和p53正常功能的突变的共存可能表明，每个突变都赋予细胞相加的生长优势。较少