Phosphoinositide-specific inositol polyphosphate 5-phosphatase IV inhibits Akt/protein kinase B phosphorylation and leads to apoptotic cell death
Phosphoinositide-specific inositol polyphosphate 5-phosphatase IV inhibits Akt/protein kinase B phosphorylation and leads to apoptotic cell death
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DOI：
10.1074/jbc.m105969200
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发表时间：
2002-02-22
影响因子：
4.8
通讯作者：
Majerus, PW
中科院分区：
文献类型：
--
作者：
Kisseleva, MV;Cao, L;Majerus, PW
磷酸肌醇特异性肌醇多磷酸5-磷酸酶IV对PI（3，4，5）P-3（Km = 0.65 μ M）具有亲和力，其比使用该底物的其它肌醇多磷酸5-磷酸酶（包括SHIP、OCRL和5 ptase II）大约10倍，这表明其在控制该代谢物的细胞内水平中可能是重要的。我们建立了稳定表达该酶的细胞系，以研究其对细胞功能的影响。我们发现，在293细胞中过表达5 ptase IV引起细胞中PI（4，5）P-2和PI（3，4，5）P-3的快速消耗，产物PI（4）P和PI（3，4）P-2相应增加，改变了细胞中磷酸肌醇3-激酶的两种磷酸肌醇产物PI（3，4）P-2和PI（3，4，5）P-3的平衡。这些磷酸肌醇的靶标之一是丝氨酸/苏氨酸激酶Akt，其在细胞凋亡的控制中起重要作用。我们能够解决PI（3，4）P-2和PI（3，4，5）P-3在Akt激活中的相对作用，通过选择性消耗这些磷酸肌醇在细胞中稳定转染5 ptase IV和肌醇多磷酸4-磷酸酶（4ptase I）。在转染4ptase I的细胞中，PI（3，4）P-2水平降低，PI（3，4，5）P-3水平升高。这两种酶的表达对Akt的磷酸化有相反的作用，以响应生长因子或热休克的刺激。Akt磷酸化在表达5 ptase IV的细胞中被抑制，但在4ptase I细胞中增加，并且与PI（3，4，5）P-3的细胞内水平相关，而与PI（3，4）P-2的细胞内水平无关。在表达5 ptase IV的细胞中Akt磷酸化的抑制使它们对FAS诱导的凋亡高度敏感，而过表达4ptase I保护细胞免于凋亡。我们的研究结果将5 ptase IV作为细胞中PI 3 K/Akt通路的相关生物调节剂。
Phosphoinositide-specific inositol polyphosphate 5-phosphatase IV has the affinity for PI(3,4,5)P-3 (K-m = 0.65 muM that is approximately 10-fold greater than the other inositol polyphosphate 5-phosphatases, which use this substrate including SHIP, OCRL, and 5ptase II, suggesting that it may be important in controlling intracellular levels of this metabolite. We created cell lines stably expressing the enzyme to study its effect on cell function. We found that overexpression of 5ptase IV in 293 cells caused the rapid depletion of both PI(4,5)P-2 and PI(3,4,5)P-3 in cells with corresponding increases in the products, PI(4)P and PI(3,4)P-2,changing the balance of two phosphoinositol products of phosphoinositide 3-kinase, PI(3,4)P-2 and PI(3,4,5)P-3, in the cell. One of the targets of these phosphoinositides is the serine/threonine kinase Akt, which plays an important role in the control of apoptosis. We were able to address the relative roles of PI(3,4)P-2 and PI(3,4,5)P-3 in the activation of Akt by selective depletion of these phosphoinositides in cells stably transfected with 5ptase IV and inositol polyphosphate 4-phosphatase (4ptase I). In cells transfected with 4ptase I, the level of PI(3,4)P-2 was reduced, and PI(3,4,5)P-3 was increased. Expression of the two enzymes had the opposite effect on the phosphorylation of Akt in response to stimulation with growth factors or heat shock. Akt phosphorylation was inhibited in cells expressing 5ptase IV but increased in 4ptase I cells and correlated with the intracellular level of PI(3,4,5)P-3 and not that of PI(3,4)P-2. The inhibition of Akt phosphorylation in cells expressing 5ptase IV makes them highly susceptible to FAS-induced apoptosis, whereas overexpressing of the 4ptase I protects cells from apoptosis. Our results place 5ptase IV as a relevant biological regulator of PI3K/Akt pathway in cells.