Alexander Kamilah, Yang Hai-Su, Hinds Philip W
Department of Pathology, Harvard Medical School, Boston, MA 02115, USA.
Mol Cancer Res. 2003 Aug;1(10):716-28.
Senescent cells in which pRb is inactivated undergo apoptosis on attempted reinitiation of DNA synthesis. To further explore the cell death resulting from loss of pRb function in senescent cells, we employed a temperature-sensitive pRb mutant protein (tspRb). We found that tspRb inactivation results in rapid E2F reactivation and subsequent S-phase reentry associated with the up-regulation of E2F target gene expression and cyclin E-dependent kinase activity. Total inhibition of cyclin-dependent kinase 2 activity results in a cell cycle arrest on pRb loss and a nearly complete suppression of apoptosis. Furthermore, blocking of E2F activity with a dominant-negative DP1 inhibits S-phase reentry and cell death following tspRb inactivation. Finally, inhibition of p73 activity abolishes apoptosis but not S-phase entry on pRb inactivation, suggesting that activation of E2F in senescent cells can result in the use of p73 as a cell death effector. Interestingly, senescent cells rescued from apoptosis maintain their altered shape and express senescence-associated beta-galactosidase despite loss of pRb function. Thus, maintenance of the terminal cell cycle arrest of senescent cells requires continuous pRb-mediated inactivation of E2F activity, the reappearance of which in these irrevocably altered cells triggers a cell death program instead of an inappropriate resumption of cell cycling.
pRb失活的衰老细胞在试图重新启动DNA合成时会发生凋亡。为了进一步探究衰老细胞中pRb功能丧失导致的细胞死亡，我们使用了一种温度敏感型pRb突变蛋白（tspRb）。我们发现，tspRb失活会导致E2F迅速重新激活，随后进入S期，这与E2F靶基因表达上调和细胞周期蛋白E依赖性激酶活性有关。完全抑制细胞周期蛋白依赖性激酶2的活性会导致pRb缺失时细胞周期停滞，并几乎完全抑制凋亡。此外，用显性负性DP1阻断E2F活性可抑制tspRb失活后的S期重新进入和细胞死亡。最后，抑制p73活性可消除pRb失活时的凋亡，但不影响S期进入，这表明衰老细胞中E2F的激活可导致将p73用作细胞死亡效应因子。有趣的是，从凋亡中挽救出来的衰老细胞尽管pRb功能丧失，但仍保持其改变的形态并表达衰老相关的β-半乳糖苷酶。因此，衰老细胞终末细胞周期停滞的维持需要pRb持续介导的E2F活性失活，而在这些不可逆转地改变的细胞中E2F活性的重新出现会触发细胞死亡程序，而不是不恰当地重新开始细胞周期。
