Dannenberg J H, van Rossum A, Schuijff L, te Riele H
The Netherlands Cancer Institute, Division of Molecular Biology, 1066 CX Amsterdam, The Netherlands.
Genes Dev. 2000 Dec 1;14(23):3051-64. doi: 10.1101/gad.847700.
The retinoblastoma suppressor pRB belongs to the family of so-called pocket proteins, which also includes p107 and p130. These proteins may functionally overlap in cell cycle control and tumor suppression. We have generated an isogenic set of embryonic stem (ES) cell lines carrying single or compound loss-of-function mutations in the Rb gene family, including a cell line completely devoid of all three pocket proteins. None of the knockout combinations affected the growth characteristics of ES cells; however, concomitant ablation of all three pocket proteins strongly impaired their differentiation capacity. For the generated genotypes, primary mouse embryonic fibroblasts (MEFs) also were obtained. While inactivation of Rb alone did not alleviate the senescence response of MEFs, pRB/p107-deficient MEFs, after having adapted to in vitro culturing, continued to proliferate at modest rate. Additional ablation of p130 rendered MEFs completely insensitive to senescence-inducing signals and strongly increased their proliferation rate. Although triple-knockout MEFs retained anchorage dependence, they lacked proper G(1) control and showed increased cell turnover under growth-inhibiting conditions.
视网膜母细胞瘤抑制蛋白pRB属于所谓的口袋蛋白家族，该家族还包括p107和p130。这些蛋白在细胞周期调控和肿瘤抑制方面可能存在功能重叠。我们构建了一组同基因的胚胎干细胞（ES）系，这些细胞系在Rb基因家族中携带单功能或复合功能丧失突变，其中包括一个完全缺失所有三种口袋蛋白的细胞系。没有一种基因敲除组合影响ES细胞的生长特性；然而，同时缺失所有三种口袋蛋白会严重损害它们的分化能力。对于所产生的基因型，还获得了原代小鼠胚胎成纤维细胞（MEF）。虽然单独失活Rb并没有减轻MEF的衰老反应，但pRB/p107缺陷型MEF在适应体外培养后，仍以适度的速率继续增殖。进一步缺失p130使MEF对衰老诱导信号完全不敏感，并显著提高了它们的增殖速率。尽管三基因敲除的MEF保留了贴壁依赖性，但它们缺乏适当的G(1)期调控，并且在生长抑制条件下细胞更新增加。
