THE HUMAN HA-RAS ONCOGENE INDUCES GENOMIC INSTABILITY IN MURINE FIBROBLASTS WITHIN ONE-CELL CYCLE
THE HUMAN HA-RAS ONCOGENE INDUCES GENOMIC INSTABILITY IN MURINE FIBROBLASTS WITHIN ONE-CELL CYCLE
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DOI：
10.1073/pnas.91.11.5124
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发表时间：
1994-05-24
影响因子：
11.1
通讯作者：
STAMBROOK, PJ
中科院分区：
文献类型：
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作者：
DENKO, NC;GIACCIA, AJ;STAMBROOK, PJ
许多人类肿瘤在ras原癌基因之一中含有激活突变。此外，这些肿瘤细胞通常是异倍体，其特征在于染色体断裂和重排，这是基因组不稳定性的结果，被认为有助于肿瘤进展。ras突变和肿瘤基因组不稳定性的同时发生促使我们问，选择性诱导激活的Ha-ras基因是否会使基因组不稳定。在这项研究中使用的NIH 3 T3细胞含有突变型p53基因，并携带选择性诱导激活（EJ）的Ha-ras转基因的控制下的细菌乳糖调节元件。当稳定转染的细胞诱导表达激活的Ha-ras异丙基β-D-硫代半乳糖苷管理，有一个显着增加的总染色体畸变的数量，包括无着丝粒片段，多着丝粒染色体，和双分钟，这发生在一个单一的细胞周期的时间范围内从诱导的时间。为了证实这些畸变发生在突变Ha-ras诱导后的第一个细胞周期内，通过血清耗竭将细胞阻滞在G（1）期，随后通过给予异丙基β-D-硫代半乳糖苷或血清将其释放。与用血清释放的平行细胞培养物的有丝分裂相比，用异丙基β-D-硫代半乳糖苷释放的细胞的有丝分裂中含有畸变的染色体部分升高3倍。因此，在这些细胞中激活的Ha-ras基因表达的诱导导致基因组的不稳定性，其可以在下一次有丝分裂时被检测为异常染色体。
Many human tumors contain an activating mutation in one of the ras protooncogenes. Additionally, these tumor cells are often heteroploid and characterized by chromosome breaks and rearrangements that are consequences of the genomic instability that is thought to contribute to tumor progression. The concurrence of ras mutations and genomic instability in tumors prompted us to ask whether selective induction of an activated Ha-ras gene could render a genome unstable. The NIH 3T3 cells used in this study contained mutant p53 genes and carried a selectively inducible activated (EJ) Ha-ras transgene under the control of bacterial lactose regulatory elements. When stably transfected cells were induced to express activated Ha-ras by isopropyl beta-D-thiogalactoside administration, there was a marked increase in the number of gross chromosomal aberrations including acentric fragments, multicentric chromosomes, and double minutes, which occurred within the time frame of a single cell cycle from the time of induction. To confirm that these aberrations occurred within the first cell cycle after mutant Ha-ras induction, the cells were arrested in G(1) phase by serum depletion and, subsequently, released by administration of isopropyl beta-D-thiogalactoside or serum. The mitoses from cells released with isopropyl beta-D-thiogalactoside contained a 3-fold elevation in the fraction of chromosomes containing aberrations compared to mitoses from parallel cell cultures that were released with serum. Thus, the induction of activated Ha-ras gene expression in these cells results in genomic instability that can be detected as aberrant chromosomes at the next mitosis.