Glut4 storage vesicles without glut4: Transcriptional regulation of insulin-dependent vesicular traffic
Glut4 storage vesicles without glut4: Transcriptional regulation of insulin-dependent vesicular traffic
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DOI：
10.1128/mcb.24.16.7151-7162.2004
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发表时间：
2004-08-01
影响因子：
5.3
通讯作者：
Pilch, PF
中科院分区：
文献类型：
--
作者：
Gross, DN;Farmer, SR;Pilch, PF
在脂肪细胞发育中起主要作用的两个转录因子家族是CCAAT/增强子结合蛋白(C/EBPs)和过氧化物酶体增殖物激活受体(PPAR)，尤其是PPARGamma。在NIH3T3成纤维细胞中异位表达C/EBPalpha或PPARGamma导致这些细胞转化为充满脂肪滴的脂肪细胞样细胞。异位表达C/EBPalpha(NIH-C/EBPalpha)的NIH 3T3细胞分化为脂肪细胞并表现出胰岛素刺激的葡萄糖摄取，而异位表达PPARGamma(NIH-PPARGamma)的NIH 3T3细胞分化但不表现出任何胰岛素刺激的葡萄糖摄取，也不表达任何C/EBPalpha。NIH-PPARGamma细胞缺乏胰岛素反应性葡萄糖摄取的原因是它们实际上缺乏胰岛素反应性葡萄糖转运体GLUT4。NIH-PPARγ细胞表达胰岛素受体-信号通路的功能活性成分(胰岛素受体、IRS-1、磷脂酰肌醇3-激酶和Akt2)，其水平与反应细胞系相当。它们还表达胰岛素敏感的囊泡运输机制的组成部分，即VAMP2、Synaxin-4和IRAP，其中最后一个是胰岛素调节的囊泡运输的另一个标志，与GLUT4一起。有趣的是，NIH-PPARγ细胞表现出正常的胰岛素依赖的IRAP易位，并分别通过细胞表面生物素化和蔗糖速度梯度分析评估形成胰岛素响应的囊泡室。此外，免疫荧光和Western印迹分析表明，GLUT4-myc结构在NIH-PPAR伽马细胞中的表达导致其胰岛素依赖的移位到质膜上。根据这些数据，我们得出结论，C/EBPalpha在NIH-PPAR伽马细胞中的主要作用是调节GLUT4的表达。分化的细胞具有较大的胰岛素敏感囊泡，GLUT4可忽略不计，GLUT4易位可在该转运蛋白表达的基础上重建。
Two families of transcription factors that play a major role in the development of adipocytes are the CCAAT/enhancer-binding proteins (C/EBPs) and the peroxisome proliferator-activated receptors (PPARs), in particular PPARgamma. Ectopic expression of either C/EBPalpha or PPARgamma in NIH 3T3 fibroblasts results in the conversion of these cells to adipocyte-like cells replete with fat droplets. NIH 3T3 cells ectopically expressing C/EBPalpha (NIH-C/EBPalpha) differentiate into adipocytes and exhibit insulin-stimulated glucose uptake, whereas NIH 3T3 cells ectopically expressing PPARgamma (NIH-PPARgamma) differentiate but do not exhibit any insulin-stimulated glucose uptake, nor do they express any C/EBPalpha The reason for the lack of insulin-responsive glucose uptake in the NIH-PPARgamma cells is their virtual lack of the insulin-responsive glucose transporter, Glut4. The NIH-PPARgamma cells express functionally active components of the insulin receptor-signaling pathway (the insulin receptor, IRS-1, phosphatidylinositol 3-kinase, and Akt2) at levels comparable to those in responsive cell lines. They also express components of the insulin-sensitive vesicular transport machinery, namely, VAMP2, syntaxin-4, and IRAP, the last of these being the other marker of insulin-regulated vesicular traffic along with Glut4. Interestingly, the NIH-PPARgamma cells show normal insulin-dependent translocation of IRAP and form an insulin-responsive vesicular compartment as assessed by cell surface biotinylation and sucrose velocity gradient analysis, respectively. Moreover, expression of a Glut4-myc construct in the NIH-PPARgamma cells results in its insulin-dependent translocation to the plasma membrane as assessed by immunofluorescence and Western blot analysis. Based on these data, we conclude that major role of C/EBPalpha in the context of the NIH-PPARgamma cells is to regulate Glut4 expression. The differentiated cells possess a large insulin-sensitive vesicular compartment with negligible Glut4, and Glut4 translocation can be reconstituted on expression of this transporter.