Caveolae and caveolin-1 are implicated in 1α,25(OH)2-vitamin D3-dependent modulation of Src, MAPK cascades and VDR localization in skeletal muscle cells
Caveolae and caveolin-1 are implicated in 1α,25(OH)2-vitamin D3-dependent modulation of Src, MAPK cascades and VDR localization in skeletal muscle cells
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DOI：
10.1016/j.jsbmb.2010.03.002
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发表时间：
2010-07-01
影响因子：
4.1
通讯作者：
Boland, Ricardo
中科院分区：
文献类型：
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作者：
Buitrago, Claudia;Boland, Ricardo
我们先前报道了1 α，25（OH）（2）D-3通过激活C2 C12骨骼肌细胞中的MAPK诱导非转录快速反应。然而，关于通过该途径启动1 α，25（OH）（2）D-3信号传导的分子机制的信息很少。质膜成分参与了一些非基因组效应。在这项工作中，我们研究了caveolae和caveolin-1（cav-1）在1 α，25（OH）（2）D-3-刺激c-Src和MAPK中的作用。当用甲基β环糊精（M β CD）（一种小窝破坏剂）预处理增殖细胞时，在细胞形态不受影响且未观察到凋亡迹象的条件下，ERK 1/2、p38 MAPK和c-Src的1 α，25（OH）（2）D-3依赖性活化受到抑制。通过siRNA技术获得了类似的结果，其中cav-1表达的沉默消除了由激素诱导的c-Src和MAPK的激活。通过共聚焦免疫细胞化学观察到cav-1与c-Src在基础条件下共定位于质膜周区。激素处理破坏了这些蛋白质的共定位，并将它们重新分配到细胞质和细胞核中。免疫共沉淀试验证实了这些观察结果。还研究了1 α，25（OH）（2）D-3暴露后VDR定位的变化。共聚焦显微镜图像显示，激素诱导VDR易位到质膜，这种作用被M β CD消除。总之，这些数据表明，小窝参与上游c-Src-MAPK激活1 α，25（OH）（2）D-3和VDR和cav-1参与快速信号由激素引起的。(C)2010爱思唯尔有限公司版权所有。
We previously reported that 1 alpha,25(OH)(2)D-3 induces non-transcriptional rapid responses through activation of MAPKs in C2C12 skeletal muscle cells. However, there is little information on the molecular mechanism underlying the initiation of 1 alpha,25(OH)(2)D-3 signaling through this pathway. Plasma membrane components have been involved in some non-genomic effects. In this work, we investigated the role of caveolae and caveolin-1 (cav-1) in 1 alpha,25(OH)(2)D-3-stimulation of c-Src and MAPKs. When proliferating cells were pretreated with methyl beta cyclodextrin (M beta CD), a caveolae disrupting agent, under conditions in which cell morphology is not affected and no signs of apoptosis are observed, 1 alpha,25(OH)(2)D-3-dependent activation of ERK1/2, p38 MAPK and c-Src was suppressed. Similar results were obtained by siRNA technology whereby silencing of cav-1 expression abolished activation of c-Src and MAPKs induced by the hormone. By confocal immunocytochemistry it was observed that cav-1 colocalizes with c-Src in the periplasma membrane zone at basal conditions. Hormone treatment disrupted the colocalization of these proteins and redistributed them into cytoplasm and nucleus. Co-immunoprecipitation assays corroborated these observations. Changes in VDR localization after 1 alpha,25(OH)(2)D-3 exposure were also investigated. Confocal microscopy images showed that the hormone induces VDR translocation to the plasma membrane, and this effect is abolished by M beta CD. Altogether, these data suggest that caveolae is involved upstream in c-Src-MAPKs activation by 1 alpha,25(OH)(2)D-3 and that VDR and cav-1 participate in the rapid signaling elicited by the hormone. (C) 2010 Elsevier Ltd. All rights reserved.