mTORC1 signaling can regulate growth factor activation of p44/42 mitogen-activated protein kinases through protein phosphatase 2A
mTORC1 signaling can regulate growth factor activation of p44/42 mitogen-activated protein kinases through protein phosphatase 2A
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DOI：
10.1074/jbc.m706173200
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发表时间：
2008-02-01
影响因子：
4.8
通讯作者：
Houghton, Peter J.
中科院分区：
文献类型：
--
作者：
Harwood, Franklin C.;Shu, Lili;Houghton, Peter J.
MTORC1复合体(雷帕霉素(MTOR)-Raptor的哺乳动物靶标)通过结节性硬化症复合体的磷酸化和失活，受到丝裂原激活蛋白(p44/42 MAP)激酶(p44/42)的调节。然而，mTORC1信号在调节p44/42激活中的作用尚未见报道。我们发现，在两个癌细胞系中，p44/42 MAPKs的调节是mTORC1依赖的。在Rh1细胞中，雷帕霉素抑制胰岛素样生长因子-I(IGF-I)刺激的Thr(202)的磷酸化，但不抑制Tyr(204)的磷酸化，并抑制p44/42激酶活性的激活。抑制mTORC1信号转导的Raptor的下调在阻断IGF-I刺激的Tyr204磷酸化方面与雷帕霉素具有相似的作用。雷帕霉素不阻断Tyr204的最大磷酸化，但延缓IGF-I刺激后Tyr204的去磷酸化速率。IGF-I对MEK1磷酸化的刺激作用(Ser217/221)不受雷帕霉素的抑制。需要较高浓度的雷帕霉素(~gt；=100 ng/ml)来抑制表皮生长因子(EGF)诱导的p44/42(Thr202)的磷酸化。低浓度冈田酸可逆转雷帕霉素对IGF-I诱导的p44/42(Thr202)磷酸化的抑制作用，提示蛋白磷酸酶2A(PP2A)参与其中。IGF-I和EGF均可引起PP2A催化亚单位(PP2Ac)从P42解离。低浓度的雷帕霉素(1 ng/ml)抑制IGF-I刺激后PP2Ac的解离，但需要较高的浓度(100 ng/ml)才能阻断EGF诱导的解离，这与雷帕霉素减弱生长因子诱导的p44/42激活的能力一致。用氨基酸剥夺法观察雷帕霉素对P44/42细胞胰岛素样生长因子-I或胰岛素活化的影响。在表达雷帕霉素耐药mTOR的Rh1mTORrr细胞中，雷帕霉素改变p44/42磷酸化动力学的作用被完全取消，而氨基酸剥夺对Rh1和Rh1mTORrr细胞的影响相似。这些结果表明，mTORC1下游的磷酸酶对p44/42的调节是复杂的。这表明mTORC1通过直接或间接调节PP2Ac来调节P44/42 MAPK上Thr202的磷酸化。
The mTORC1 complex ( mammalian target of rapamycin (mTOR)-raptor) is modulated by mitogen-activated protein (p44/42 MAP) kinases (p44/42) through phosphorylation and inactivation of the tuberous sclerosis complex. However, a role for mTORC1 signaling in modulating activation of p44/42 has not been reported. We show that in two cancer cell lines regulation of the p44/42 MAPKs is mTORC1-dependent. In Rh1 cells rapamycin inhibited insulin-like growth factor-I (IGF-I)-stimulated phosphorylation of Thr(202) but not Tyr(204) and suppressed activation of p44/42 kinase activity. Down-regulation of raptor, which inhibits mTORC1 signaling, had a similar effect to rapamycin in blocking IGF-I- stimulated Tyr204 phosphorylation. Rapamycin did not block maximal phosphorylation of Tyr204 but retarded the rate of dephosphorylation of Tyr204 following IGF-I stimulation. IGF-I stimulation of MEK1 phosphorylation (Ser(217/221)) was not inhibited by rapamycin. Higher concentrations of rapamycin (>= 100 ng/ml) were required to inhibit epidermal growth factor (EGF)induced phosphorylation of p44/42 ( Thr202). Rapamycin-induced inhibition of p44/42 ( Thr202) phosphorylation by IGF-I was reversed by low concentrations of okadaic acid, suggesting involvement of protein phosphatase 2A(PP2A). Both IGF-I and EGF caused dissociation of PP2A catalytic subunit (PP2Ac) from p42. Whereas low concentrations of rapamycin ( 1 ng/ml) inhibited dissociation of PP2Ac after IGF-I stimulation, it required higher concentrations (> 100 ng/ ml) to block EGF-induced dissociation, consistent with the ability for rapamycin to attenuate growth factor-induced activation of p44/42. The effect of rapamycin on IGF-I or insulin activation of p44/42 was recapitulated by amino acid deprivation. Rapamycin effects altering the kinetics of p44/42 phosphorylation were completely abrogated in Rh1mTORrr cells that express a rapamycin-resistant mTOR, whereas the effects of amino acid deprivation were similar in Rh1 and Rh1mTORrr cells. These results indicate complex regulation of p44/42 by phosphatases downstream of mTORC1. This suggests a model in which mTORC1 modulates the phosphorylation of Thr202 on p44/42 MAPKs through direct or indirect regulation of PP2Ac.