Okoumassoun Liliane Eustache, Russo Caterina, Denizeau Francine, Averill-Bates Diana, Henderson Janet E
JTN Wong Laboratories for Mineralized Tissue Research, McGill University, Montreal, Québec, Canada.
J Cell Physiol. 2007 Sep;212(3):591-9. doi: 10.1002/jcp.21055.
Over the past decade, parathyroid hormone-related protein (PTHrP) has been identified as a key survival factor for cells subjected to apoptotic stimuli. Its anti-apoptotic activity has been attributed to nuclear accumulation of the intact protein, or a synthetic peptide corresponding to its nuclear targeting sequence (NTS), which promotes rapid exit of nutrient deprived cells from the cell cycle. Intracellular PTHrP also inhibited apoptosis by blocking tumor necrosis factor alpha (TNFalpha)-induced apoptosis by blocking signaling from the "death receptor" and preventing damage to the mitochondrial membrane. In both cases, the anti-apoptotic activity was significantly reduced in the presence of a nuclear deficient form of PTHrP with a (88)K/E K/E.K/I(91) mutation in the NTS. The current work was undertaken to determine the mechanism by which nuclear PTHrP blocked mitochondrial-mediated apoptosis. Using sub-cellular fractionation and functional assays we showed that pre-treatment of HEK293 cells with exogenous NTS peptide before inducing apoptosis with TNFalpha was as effective as expression of the full-length protein in inhibiting apoptosis. Inhibition of apoptosis was associated with increased expression of protein kinase casein kinase 2 (CK2) and in sustained CK2 accumulation and activity in the nuclear fraction. In primary chondrogenic cells harvested from the limb buds of PTHrP(+/-) and PTHrP(-/-) embryonic mice, there was a dose-dependent decrease in CK2 expression and activity that correlated with increased susceptibility to apoptosis. Taken together the results indicate that nuclear accumulation of PTHrP effectively inhibits mitochondrial-mediated apoptosis through regulation of the expression, activity, and sub-cellular trafficking of CK2.
在过去十年中，甲状旁腺激素相关蛋白（PTHrP）已被确定为遭受凋亡刺激的细胞的关键生存因子。其抗凋亡活性归因于完整蛋白或与其核靶向序列（NTS）对应的合成肽在细胞核内的积累，这促进营养缺乏的细胞快速退出细胞周期。细胞内的PTHrP还通过阻断肿瘤坏死因子α（TNFα）诱导的凋亡来抑制凋亡，即通过阻断“死亡受体”的信号传导并防止线粒体膜受损。在这两种情况下，当存在NTS中具有（88）K/E K/E.K/I（91）突变的核缺陷形式的PTHrP时，抗凋亡活性显著降低。目前的研究旨在确定细胞核内PTHrP阻断线粒体介导的凋亡的机制。通过亚细胞分级分离和功能测定，我们发现用外源性NTS肽预处理HEK293细胞，然后用TNFα诱导凋亡，在抑制凋亡方面与全长蛋白的表达效果相同。凋亡的抑制与蛋白激酶酪蛋白激酶2（CK2）表达增加以及CK2在细胞核部分的持续积累和活性有关。在从PTHrP（+/-）和PTHrP（-/-）胚胎小鼠的肢芽中收获的原代软骨细胞中，CK2的表达和活性呈剂量依赖性降低，这与对凋亡的易感性增加相关。综合这些结果表明，PTHrP在细胞核内的积累通过调节CK2的表达、活性和亚细胞转运有效地抑制线粒体介导的凋亡。
