Bi-directional regulation of Ser-985 phosphorylation of c-Met via protein kinase C and protein phosphatase 2A involves c-Met activation and cellular responsiveness to hepatocyte growth factor
Bi-directional regulation of Ser-985 phosphorylation of c-Met via protein kinase C and protein phosphatase 2A involves c-Met activation and cellular responsiveness to hepatocyte growth factor
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DOI：
10.1074/jbc.m314254200
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发表时间：
2004-06-18
影响因子：
4.8
通讯作者：
Nakamura, T
中科院分区：
文献类型：
--
作者：
Hashigasako, A;Machide, M;Nakamura, T
以前的研究表明，用12-O-十四酰佛波醇-13-乙酸酯处理细胞诱导c-Met（肝细胞生长因子（HGF）的受体酪氨酸激酶）胞膜上Ser-985的磷酸化，并且这与c-Met的酪氨酸磷酸化降低有关。然而，c-Met中Ser-985磷酸化的调节机制和生物学意义仍然未知。当A549人肺癌细胞暴露于氧化应激与H2 O2，H2 O2处理诱导磷酸化的Ser-985，但这是废除蛋白激酶C（PKC）的抑制剂。同样地，用NaF（蛋白磷酸酶的抑制剂）处理细胞允许Ser-985磷酸化，并且负责Ser-985去磷酸化的蛋白磷酸酶被鉴定为蛋白磷酸酶2A（PP 2A）。pull-down分析表明，PKC δ和PKC β与c-Met的Ser-985磷酸化有关，PKC δ和PKC β可能参与c-Met的Ser-985磷酸化调节。相反，PP 2A与c-Met组成性相关，而当细胞暴露于H2 O2时，其使c-Met的Ser-985去磷酸化的活性降低。在培养的A549细胞中加入HGF诱导c-Met酪氨酸磷酸化，结果是促有丝分裂反应和细胞分散。相反，在H2 O2应激的存在下，HGF依赖的c-Met酪氨酸磷酸化在很大程度上被抑制，与Ser-985磷酸化呈相互关系，并且该事件与细胞对HGF的反应性的消除相关。这些结果表明c-Met的Ser-985磷酸化通过PKC和PP 2A双向调节，并且Ser-985磷酸化状态可能提供了一种独特的机制，该机制赋予细胞对HGF的反应性/非反应性，这取决于细胞外条件。
Previous studies indicated that treatment of cells with 12-O-tetradecanoylphorbol-13-acetate induced phosphorylation of Ser-985 at the juxtamembrane of c-Met, the receptor tyrosine kinase for hepatocyte growth factor (HGF), and this was associated with decreased tyrosine phosphorylation of c-Met. However, the regulatory mechanisms and the biological significance of the Ser-985 phosphorylation in c-Met remain unknown. When A549 human lung cancer cells were exposed to oxidative stress with H2O2, H2O2 treatment induced phosphorylation of Ser-985, but this was abrogated by an inhibitor for protein kinase C (PKC). Likewise, treatment of cells with NaF (an inhibitor of protein phosphatases) allowed for phosphorylation of Ser-985, and a protein phosphatase responsible for dephosphorylation of Ser-985 was identified to be protein phosphatase 2A (PP2A). The effects of PKC inhibitors revealed that PKCdelta and -epsilon were responsible for the Ser-985 phosphorylation of c-Met, and pull-down analysis indicated that associations of PKCdelta and -epsilon with c-Met may be involved in the regulation of Ser-985 phosphorylation of c-Met. Instead, PP2A was constitutively associated with c-Met, whereas its activity to dephosphorylate Ser-985 of c-Met was decreased when cells were exposed to H2O2. Addition of HGF to A549 cells in culture induced c-Met tyrosine phosphorylation, the result being mitogenic response and cell scattering. In contrast, in the presence of H2O2 stress, HGF-dependent tyrosine phosphorylation of c-Met was largely suppressed with a reciprocal relationship to Ser-985 phosphorylation, and this event was associated with abrogation of cellular responsiveness to HGF. These results indicate that Ser-985 phosphorylation of c-Met is bi-directionally regulated through PKC and PP2A, and the Ser-985 phosphorylation status may provide a unique mechanism that confers cellular responsiveness/unresponsivenss to HGF, depending on extracellular conditions.