Intersectin regulates fission and internalization of caveolae in endothelial cells.
Intersectin regulates fission and internalization of caveolae in endothelial cells.
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Intersectin 调节内皮细胞中小窝的裂变和内化。
DOI：
10.1091/mbc.e03-01-0041
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发表时间：
2003
影响因子：
3.3
通讯作者：
Malik,AsrarB
中科院分区：
文献类型：
--
作者：
Predescu,SandaA;Predescu,DanN;Timblin,BarbaraK;Stan,RaduV;Malik,AsrarB
Intersectin是一种含有多种Eps 15同源性和Src同源性3（SH 3）结构域的蛋白，是神经元和非神经元细胞内吞机制的组成部分。然而，其在内皮细胞（EC）中通过小窝的内吞作用中的作用尚不清楚。我们在此证明，通过共免疫沉淀，速度沉降甘油梯度，和交联的intersectin是目前在EC中的膜相关蛋白复合物，含有发动蛋白和SNAP-23。电子显微镜（EM）的免疫金标记研究表明，interstin优先与小窝颈部，它仍然与小窝后，从质膜分裂。一个无细胞系统耗尽intersectin未能支持小窝裂变从质膜。用于定量小窝内化的生物素测定和过表达wt-交叉蛋白的EC的广泛EM形态学分析表明了广泛的形态学变化（即，大的小窝簇在细胞周边和多形小窝颈处边缘化）以及受损的小窝内化。通过ELISA对小窝介导的摄取进行的生物化学评价显示，相对于对照，抑制率为68.4%。我们还表明，intersectin与发动蛋白的相互作用是重要的，在调节细胞膜小窝的分裂和内化。两者合计，结果表明，在内皮细胞中的小窝分裂的机制中的关键作用的intersectin。
Intersectin, a multiple Eps15 homology and Src homology 3 (SH3) domain–containing protein, is a component of the endocytic machinery in neurons and nonneuronal cells. However, its role in endocytosis via caveolae in endothelial cells (ECs) is unclear. We demonstrate herein by coimmunoprecipitation, velocity sedimentation on glycerol gradients, and cross-linking that intersectin is present in ECs in a membrane-associated protein complex containing dynamin and SNAP-23. Electron microscopy (EM) immunogold labeling studies indicated that intersectin associated preferentially with the caveolar necks, and it remained associated with caveolae after their fission from the plasmalemma. A cell-free system depleted of intersectin failed to support caveolae fission from the plasma membrane. A biotin assay used to quantify caveolae internalization and extensive EM morphological analysis of ECs overexpressing wt-intersectin indicated a wide range of morphological changes (i.e., large caveolae clusters marginated at cell periphery and pleiomorphic caveolar necks) as well as impaired caveolae internalization. Biochemical evaluation of caveolae-mediated uptake by ELISA showed a 68.4% inhibition by reference to control. We also showed that intersectin interaction with dynamin was important in regulating the fission and internalization of caveolae. Taken together, the results indicate the crucial role of intersectin in the mechanism of caveolae fission in endothelial cells.
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细胞表面生物素化技术
DOI：
10.1016/b978-0-12-164717-9.50028-3
发表时间：
1994
期刊：
The journal of physical chemistry. A
影响因子：
--
作者：
C. Zurzolo;A. Bivic;E. Rodriguez
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DOI：
--
发表时间：
1991
期刊：
Cell biology reviews : CBR
影响因子：
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作者：
Simionescu,M;Simionescu,N
通讯作者：
Simionescu,N
影响因子：
3.5
作者：
Pucharcos, C;Estivill, X;de la Luna, S
通讯作者：
de la Luna, S
影响因子：
7.5
作者：
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影响因子：
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作者：
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