p300 is required for orderly G1/S transition in human cancer cells
p300 is required for orderly G1/S transition in human cancer cells
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DOI：
10.1038/sj.onc.1209771
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发表时间：
2007-01-04
期刊：
影响因子：
8
通讯作者：
Caldas, C.
中科院分区：
文献类型：
--
作者：
Iyer, N. G.;Xian, J.;Caldas, C.
由于缺乏合适的实验系统，转录共激活因子p300在细胞周期调控中的作用尚未得到详细分析。我们现在已经检测了p300缺陷型癌细胞系的细胞周期进程，在这些细胞系中，p300要么通过基因打靶（p300（-）细胞）被破坏，要么使用RNA干扰被敲低。尽管在正常生长条件下存在显著的增殖缺陷，但p300（-）缺陷型细胞迅速通过G1期并过早进入S期。加速的G1/S转换与早期视网膜母细胞瘤（RB）过度磷酸化以及E2F靶标的激活有关。p300（-）的乙酰转移酶活性并非必需，因为一个组蛋白乙酰转移酶（HAT）缺陷型p300突变体的表达逆转了这些变化。免疫共沉淀显示p300/RB相互作用在G1期体内发生，并且这种相互作用有两个峰值：在G1早期与未磷酸化的RB，以及在G1晚期与磷酸化的RB。体外激酶实验表明p300直接抑制cdk6介导的RB磷酸化，这表明p300在G1早期起作用以防止RB过度磷酸化并延迟过早进入S期。矛盾的是，观察到尽管血清耗竭时间延长，p300（-）细胞仍持续循环，并且这与持续的RB过度磷酸化有关。总之，这些结果表明p300在G1/S调控中具有重要作用，可能是通过调节RB磷酸化来实现的。
The role of the transcriptional coactivator p300 in cell cycle control has not been analysed in detail due to the lack of appropriate experimental systems. We have now examined cell cycle progression of p300-deficient cancer cell lines, where p300 was disrupted either by gene targeting (p300(-) cells) or knocked down using RNAi. Despite significant proliferation defects under normal growth conditions, p300(-) deficient cells progressed rapidly through G1 with premature S-phase entry. Accelerated G1/S transition was associated with early retinoblastoma (RB) hyperphosphorylation and activation of E2F targets. The p300(-) acetylase activity was dispensable since expression of a HAT-deficient p300 mutant reversed these changes. Co-immunoprecipitation showed p300/RB interaction occurs in vivo during G1, and this interaction has two peaks: in early G1 with unphosphorylated RB and in late G1 with phosphorylated RB. In vitro kinase assays showed that p300 directly inhibits cdk6-mediated RB phosphorylation, suggesting p300 acts in early G1 to prevent RB hyperphosphorylation and delay premature S-phase entry. Paradoxically, continued cycling of p300(-) cells despite prolonged serum depletion was observed, and this occurred in association with persistent RB hyperphosphorylation. Altogether, these results suggest that p300 has an important role in G1/S control, possibly by modulating RB phosphorylation.