Gabai Vladimir L, O'Callaghan-Sunol Cornelia, Meng Le, Sherman Michael Y, Yaglom Julia
Department of Biochemistry, Boston University Medical School, Boston, MA 02118, USA.
Cancer Res. 2008 Mar 15;68(6):1834-42. doi: 10.1158/0008-5472.CAN-07-5656.
Depletion of the major heat shock protein Hsp72 leads to activation of the senescence program in a variety of tumor cell lines via both p53-dependent and p53-independent pathways. Here, we found that the Hsp72-depleted cells show defect in phosphorylation and activation of the protein kinase Chk1 by genotoxic stresses, such as UVC irradiation or camptothecin. Under these conditions, phosphorylation of Rad17 was also suppressed, whereas phosphorylation of p53 at Ser(15) was not affected, indicating a specific defect in phosphorylation of a subset of the ATR kinase substrates. Similarly, suppression of Chk1 activation was seen when senescence signaling was triggered by direct stimulation of p53, depletion of Cdc2, or overexpression of the cell cycle inhibitors p21 or p16. Thus, defect in Chk1 activation was not a consequence of the chaperone imbalance, but rather a downstream effect of activation of the senescence signaling. Inhibition of Chk1 was associated with inefficient inter-S phase checkpoint, as Hsp72 depleted cells failed to halt cell cycle progression upon UVC irradiation. Accordingly, sensitivity of cells to genotoxic stimuli after Hsp72 depletion was significantly enhanced. Thus, activation of the senescence signaling causes a defect in the DNA damage response manifested in increased sensitivity to genotoxic stresses.
主要热休克蛋白Hsp72的缺失通过p53依赖和p53非依赖途径导致多种肿瘤细胞系中衰老程序的激活。在此，我们发现Hsp72缺失的细胞在受到UVC照射或喜树碱等基因毒性应激时，蛋白激酶Chk1的磷酸化和激活存在缺陷。在这些条件下，Rad17的磷酸化也受到抑制，而p53在Ser(15)位点的磷酸化不受影响，这表明ATR激酶底物的一个子集在磷酸化方面存在特异性缺陷。同样，当通过直接刺激p53、耗尽Cdc2或过表达细胞周期抑制剂p21或p16触发衰老信号时，也观察到Chk1激活受到抑制。因此，Chk1激活缺陷不是伴侣蛋白失衡的结果，而是衰老信号激活的下游效应。Chk1的抑制与S期内检查点功能低下有关，因为Hsp72缺失的细胞在UVC照射后未能阻止细胞周期进程。相应地，Hsp72缺失后细胞对基因毒性刺激的敏感性显著增强。因此，衰老信号的激活导致DNA损伤反应缺陷，表现为对基因毒性应激的敏感性增加。
