Mechanistic Analysis of a DNA Damage-Induced, PTEN-Dependent Size Checkpoint in Human Cells
Mechanistic Analysis of a DNA Damage-Induced, PTEN-Dependent Size Checkpoint in Human Cells
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DOI：
10.1128/mcb.01323-10
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发表时间：
2011-07-01
影响因子：
5.3
通讯作者：
Waldman, Todd
中科院分区：
文献类型：
--
作者：
Kim, Jung-Sik;Xu, Xuehua;Waldman, Todd
DNA损伤后，人类细胞在细胞周期的G(1)和G(2)阶段受阻，细胞大小同时受阻。我们先前证明，PTEN肿瘤抑制基因的突变失活可以使细胞大小受阻与细胞周期受阻解偶联。在这里，我们证明了细胞大小检查点可以被DNA损伤的化疗药物和电离辐射诱导，并受到PTEN的有效调节，但不受其致癌同行PIK3CA的调节。PTEN的突变分析和Akt的药理抑制表明，Akt的磷酸化调控对于细胞大小检查点的控制是不必要的。为了发现可能参与大小检查点控制的PTEN调节因子和/或效应因子，我们采用了一种新的内源性表位标记(EET)方法，该方法揭示了内源性PTEN与肌动蛋白重塑复合体相互作用，该复合体包括肌动蛋白、明胶蛋白和EPLIN。药物抑制PTEN(+/+)细胞肌动蛋白重塑，重现了PTEN(-/-)细胞缺乏大小检查点控制的现象。综上所述，这些结果进一步支持了DNA损伤诱导的大小检查点的存在，该检查点由主要的肿瘤抑制因子调控，并提供了一种新的Akt非依赖机制，通过该机制PTEN控制细胞大小。
Following DNA damage, human cells undergo arrests in the G(1) and G(2) phases of the cell cycle and a simultaneous arrest in cell size. We previously demonstrated that the cell size arrest can be uncoupled from the cell cycle arrest by mutational inactivation of the PTEN tumor suppressor gene. Here we show that the cell size checkpoint is inducible by DNA-damaging chemotherapeutic agents as well as by ionizing radiation and is effectively regulated by PTEN but not by its oncogenic counterpart, PIK3CA. Mutational analysis of PTEN and pharmacological inhibition of Akt revealed that modulation of Akt phosphorylation is unnecessary for cell size checkpoint control. To discover putative PTEN regulators and/or effectors involved in size checkpoint control, we employed a novel endogenous epitope tagging (EET) approach, which revealed that endogenous PTEN interacts at the membrane with an actin-remodeling complex that includes actin, gelsolin, and EPLIN. Pharmacological inhibition of actin remodeling in PTEN(+/+) cells recapitulated the lack of size checkpoint control seen in PTEN(-/-) cells. Taken together, these results provide further support for the existence of a DNA damage-inducible size checkpoint that is regulated by a major tumor suppressor, and they provide a novel Akt-independent mechanism by which PTEN controls cell size.