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Role of MAP Kinase Cascade in the Regulation of Diverse Cellular Fuctions.

MAP 激酶级联在多种细胞功能调节中的作用。

基本信息

批准号：
10470485
负责人：
KOHNO Michiaki
金额：
$ 8.45万
依托单位：
Nagasaki University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B).
财政年份：
1998
资助国家：
日本
起止时间：
1998 至 2000
项目状态：
已结题

项目摘要

(1) Blockade of the ERK-MAP kinase pathway by treatment with PD98059, a specific inhibitor of MEK, completely suppressed the growth of tumor cells in which the pathway is constitutively activated. Selective up-regulation of p27^<Klp1> was observed after PD98059 treatment of these tumor cells. The up-regulation of p27^<Klp1> correlated with increased association of p27^<Klp1> with cyclin E-CDK2 complexes, a concomitant inhibition of cyclin E-CDK2 kinase activity, and consequent decrease in the phosphorylation state of RB, which would culminate in the marked G1 cell cycle arrest observed in these tumor cells. These results suggest that the complete growth suppression that follows specific blockade of the ERK-MAP kinase pathway in tumor cells in which the pathway is constitutively activated is mediated by up-regulation of p27^<Klp1>.(2) HGF induced the spreading, dissociation and scattering of MDCK cells. HGF induced the sustained activation of ERK-MAP kinases in the cells. We have examin … More ed whether or not the ERK-MAP kinase activity in the nucleus is required for HGF-induced scattering of MDCK cells. For the analysis, we forced cytoplasmic retention of ERK-MAP kinases in the cells by expressing an inactive form of the MKP-3 cytoplasmic phosphatase. The enforced cytoplasmic retention of the activated ERK-MAP kinases apparently inhibited the scattering response of MDCK cells in response to HGF.These results indicate that the ERK-MAP kinase activity in the nucleus is required for the motility response of MDCKcells induced by HGF.(3) NGF has the capacity to induce the neuronal differentiation of PC12 cells. NGF induced the sustained activation of p38 MAP kinase pathway as well as of ERK-MAP kinase pathway. Pretreatment of PC12 cells with SB203580 (a specific inhibitor of p38 MAP kinase) or PD98059 partially inhibited the NGF-induced neurite outgrowth formation, while pretreatment of the cells with a combination of PD98059 and SB203580 resulted in almost complete inhibition of NGF-induced neurite outgrowth. These results suggest that activation of the ERK-MAP kinase pathway together with activation of p38 MAP kinase pathway are required for full induction of neurite outgrowth following stimulation of PC12 cells with NGF.ERK-MAP kinases are suggestively involved in the phosphorylation of neurofilament proteins (NFs), while p38 MAP kinase is involved in the expression of NF genes. Less

(1)通过MEK特异性抑制剂PD98059治疗阻断ERK-MAP激酶通路，完全抑制该通路被组成性激活的肿瘤细胞的生长。 PD98059处理这些肿瘤细胞后观察到p27^<Klp1>的选择性上调。 p27^<Klp1> 的上调与 p27^<Klp1> 与细胞周期蛋白 E-CDK2 复合物的关联增加相关，同时抑制细胞周期蛋白 E-CDK2 激酶活性，并随后降低 RB 的磷酸化状态，最终导致在这些肿瘤细胞中观察到的显着的 G1 细胞周期停滞。这些结果表明，在肿瘤细胞中，ERK-MAP激酶途径被特异性阻断后，该途径被组成性激活，随后的完全生长抑制是由p27^<Klp1>的上调介导的。(2)HGF诱导MDCK细胞的扩散、解离和分散。 HGF 诱导细胞中 ERK-MAP 激酶的持续激活。我们检查了细胞核中的 ERK-MAP 激酶活性是否是 HGF 诱导的 MDCK 细胞分散所必需的。为了进行分析，我们通过表达非活性形式的 MKP-3 细胞质磷酸酶来强制 ERK-MAP 激酶在细胞中保留在细胞质中。激活的ERK-MAP激酶的强制滞留明显抑制了MDCK细胞对HGF的散射反应。这些结果表明细胞核内的ERK-MAP激酶活性是HGF诱导的MDCK细胞运动反应所必需的。(3)NGF具有诱导PC12细胞神经元分化的能力。 NGF 诱导 p38 MAP 激酶途径以及 ERK-MAP 激酶途径的持续激活。用SB203580（p38 MAP激酶的特异性抑制剂）或PD98059预处理PC12细胞部分抑制NGF诱导的神经突生长形成，而用PD98059和SB203580组合预处理细胞导致几乎完全抑制NGF诱导的神经突生长。这些结果表明，用 NGF 刺激 PC12 细胞后，完全诱导神经突生长需要 ERK-MAP 激酶途径的激活和 p38 MAP 激酶途径的激活。ERK-MAP 激酶可能参与神经丝蛋白 (NF) 的磷酸化，而 p38 MAP 激酶参与 NF 基因的表达。较少的