Simultaneous phosphorylation of p53 at serine 15 and 20 induces apoptosis in human glioma cells by increasing expression of pro-apoptotic genes
Simultaneous phosphorylation of p53 at serine 15 and 20 induces apoptosis in human glioma cells by increasing expression of pro-apoptotic genes
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DOI：
10.1007/s11060-009-9844-1
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发表时间：
2009-05-01
影响因子：
3.9
通讯作者：
Lang, Frederick F.
中科院分区：
文献类型：
--
作者：
Amano, Toshiyuki;Nakamizo, Akira;Lang, Frederick F.
了解 p53 诱导细胞周期停滞与细胞凋亡的能力的机制对于治疗人类神经胶质瘤至关重要，其中 70% 的神经胶质瘤含有野生型 p53。尽管 N 末端磷酸化会导致 p53 的激活，但 N 末端磷酸化（尤其是丝氨酸 15 和 20）在 p53 诱导细胞周期停滞与细胞凋亡的能力中的作用仍然存在争议。在这里，我们检验了以下假设：丝氨酸 15 和/或 20 的磷酸化与人类神经胶质瘤中 p53 介导的细胞凋亡存在因果关系。在相同位点引入含有丝氨酸 15 或/和丝氨酸 20 丙氨酸突变（阻止磷酸化）或天冬氨酸突变（模拟磷酸化）的 p53 质粒，表明两个位点同时磷酸化可诱导细胞凋亡。当双磷酸化模拟腺病毒 p53 载体 (Ad-p53-15D20D) 与未磷酸化 p53 载体 (Ad-p53) 进行比较时，Ad-p53 处理导致 G1 期停滞，而 Ad-p53-15D20D 诱导细胞凋亡。这些作用的发生与其他 N 端丝氨酸（即丝氨酸 6、9、33、37、46）的磷酸化无关，表明 S15 和 S20 的磷酸化足以诱导细胞凋亡。从机制上讲，Ad-p53 只能增加 p21/CIP 的表达，而 Ad-p53-15D20D 则增加促凋亡基因 Fas、Puma 和 PIG3 的结合和表达。然而，Ad-p53-15D20D 并没有改变 Noxa、Bid、IGFBP3、PERP 和 Killer/DR5 的表达，表明 S15 和 S20 的磷酸化导致了特定促凋亡基因的表达。总之，S15 和 S20 同时磷酸化与细胞凋亡有因果关系，导致特定 p53 响应性促凋亡基因的表达增加。
Understanding the mechanism underlying p53's ability to induce cell cycle arrest versus apoptosis is critical to treating human gliomas, 70% of which contain wild-type p53. Although N-terminal phosphorylation results in activation of p53, the role of N-terminal phosphorylation, particularly at serines 15 and 20, in p53's ability to induce cell cycle arrest versus apoptosis remains controversial. Here we test the hypothesis that phosphorylation of serine 15 and/or 20 is causally related to p53-mediated apoptosis in human gliomas. Introduction of p53 plasmids containing alanine mutations at serine 15 or/and serine 20 (which block phosphorylation) or aspartate mutations (which mimic phosphorylation) at the same sites, implicated simultaneous phosphorylation of both sites in the induction of apoptosis. When a double phosphorylation-mimicking adenoviral p53 vector (Ad-p53-15D20D) was compared with an unphosphorylated p53 vector (Ad-p53), treatment with Ad-p53 resulted in G1-arrest, whereas Ad-p53-15D20D induced apoptosis. These effects occurred independent of phosphorylation of other N-terminal serine (i.e., serines 6, 9, 33, 37, 46) indicating that phosphorylation of S15 and S20 is sufficient for inducing apoptosis. Mechanistically, Ad-p53 was capable only of increasing the expression of p21/CIP, whereas Ad-p53-15D20D increased the binding to and expression of the pro-apoptotic genes Fas, Puma and PIG3. However, Ad-p53-15D20D did not alter the expression of Noxa, Bid, IGFBP3, PERP and Killer/DR5, suggesting that phosphorylation of S15 and S20 resulted in the expression of specific pro-apoptotic gene. In conclusion, simultaneous phosphorylation of S15 and S20 is causally associated with apoptosis, resulting in increased expression of specific p53-responsive pro-apoptotic genes.