Dynamics of nuclear shuttling with MAP kinase

MAP 激酶的核穿梭动力学

基本信息

批准号：
12480202
负责人：
NAKANISHI Mamoru
金额：
$ 4.42万
依托单位：
Nagoya City University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2002
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-12480202/
关键词：
MAP kinase MAP kinase cascade GFP Confocal microscope Fluorescent protein Raf-1 Nuclear shuttling Basophile キメラ蛋白質核シヤトル

项目摘要

The mitogen-activated protein (MAP) kinase cascade consists of MAP kinase (MAP; ERK2) and its activator, MAP kinase (MAPKK; MEK). However, the mechanisms for activation of ERK2 have not been defined yet in cells. Here, we used fluorescent-tagged ERK2 and MEK to examine the localization of ERK2 and MEK in living rat basophilic leukemia (RBL-2H3) cells. ERK2 was mainly in the cytoplasm in resting cells, but translocated into the nucleus after the ligation of IgE receptors. The import of ERK2 reached the maximum at 6-7 min, and then the imported ERK2 was exported from the nucleus. MEK mainly resided in the cytoplasm, and no significant MEK translocation was detected after ligation of IgE receptors. The data suggested that the sustained calcium increase was required for the optimal translocation of ERK2 into the nucleus in RBL cells. However, analysis of the dynamics of ERK2 and MEK suggested that both of them rapidly shuttle between the cytoplasm and the nucleus, and that MEK regulate the … More nuclear shuttling of ERK2 while MEK remain mainly in the cytoplasm. Pretreatment of RBL-2H3 cells with cytochalasin D and Latrunculin A, inhibitors of actin polymerization, prolonged the nuclear translocation of ERK2 after antigen stimulation. Western blotting analysis revealed that these drugs enhanced the phosphorylation of both ERK2 and MEK after antigen stimuation. To the contrary, nocodazole, an inhibitor of tubulin polymerization, caused neither prolongation of nuclear translocation of ERK2 nor enhancement of phosphorylation of ERK2 and MEK. Dissociation of cross-linked receptors by the addition of excess amount of monovalent hapten halted translocation of ERK2 even in cells pretreated with cytochalasin D. Taken together, these results indicate that actin polymerization affects the nuclear shuttling of ERK2 by the regulation of IgE receptor cross-linking. These results gave a new insight of the dynamics of ERK2 and MEK in the nuclear shuttling of RBL-2H3 cells after Ag stimulation. Less

有丝分裂原活化蛋白（MAP）激酶级联反应由MAP激酶（MAP; ERK2）及其激活剂MAPKK组成，用于激活ERK2的MEK; MEK检查活的大鼠嗜碱性白血病（RBL-2H3）细胞。 ERK2是从核中出口的。核，MEK调节了更多的ERK2的核穿梭，而MEK则保留在细胞质中Nocodazole是微管蛋白聚合的抑制剂，既不导致ERK2的核易位，也不导致ERK2和MEK的磷酸化的增强，通过添加过量的单交易量的交联受体分离。通过调节IgE受体交叉链接的ERK2的核梭式化