Single cell monitoring of growth arrest and morphological changes induced by transfer of wild-type p53 alleles to glioblastoma cells.
Single cell monitoring of growth arrest and morphological changes induced by transfer of wild-type p53 alleles to glioblastoma cells.
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单细胞监测野生型 p53 等位基因转移至胶质母细胞瘤细胞引起的生长停滞和形态变化。
DOI：
10.1073/pnas.92.4.1008
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发表时间：
1995
影响因子：
11.1
通讯作者：
W. Cavenee
中科院分区：
文献类型：
--
作者：
Erwin G. Van Meir;K. Roemer;A. Diserens;T. Kikuchi;S. Rempel;M. Haas;H. Huang;T. Friedmann;N. de Tribolet;W. Cavenee
p53抑癌基因突变是人类星形细胞瘤恶性进展过程中最早发现的遗传性病变之一。为了评估这些突变的功能意义，将野生型（WT）p53基因引入具有突变型、WT或无效内源性p53等位基因的胶质母细胞瘤细胞系中。在培养物中自发选择具有突变或无效内源性p53等位基因和外源性WT p53的细胞群体，用于仅表达突变型p53或不表达p53的细胞，然后其显示与亲本细胞相同的生长和致瘤表型。为了确定突变发生前WT p53表达的表型结果，我们开发了一种单细胞测定法来监测WT p53依赖性转录活性。将外源性WT p53基因转移和表达至具有内源性突变或缺失但非WT p53等位基因的细胞引起生长停滞和形态学变化，包括细胞大小增加和获得多个核。这支持了p53基因的遗传损伤在星形细胞瘤的发生中起重要作用的假设。此外，在此开发的附加型单细胞报告策略的高灵敏度在快速筛选p53基因或具有已知转录反式激活靶点的任何其他基因的生殖系突变的患者中具有潜在的临床应用。
Mutation of the p53 tumor suppressor gene is one of the earliest identified genetic lesions during malignant progression of human astrocytomas. To assess the functional significance of these mutations, wild-type (WT) p53 genes were introduced into glioblastoma cell lines having mutant, WT, or null endogenous p53 alleles. Populations of cells with mutant or null endogenous p53 alleles and exogenous WT p53 were spontaneously selected in culture for cells expressing only mutant p53 or no p53, which then displayed a growth and tumorigenic phenotype identical to the parental cells. To determine the phenotypic consequences of WT p53 expression before the occurrence of mutations, we developed a single cell assay to monitor WT p53-dependent transcription activity. Transfer and expression of exogenous WT p53 genes to cells with endogenous mutant or deleted, but not WT, p53 alleles caused growth arrest and morphological changes, including increased cell size and acquisition of multiple nuclei. This supports the hypothesis that genetic lesions of the p53 gene play an important role in the genesis of astrocytomas. Furthermore, the high sensitivity of the episomal single cell reporter strategy developed here has potential clinical applications in the rapid screening of patients for germ-line mutations of the p53 gene or any other gene with known targets for transcriptional transactivation.