Brain slice biotinylation: an ex vivo approach to measure region-specific plasma membrane protein trafficking in adult neurons.
Brain slice biotinylation: an ex vivo approach to measure region-specific plasma membrane protein trafficking in adult neurons.
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DOI：
10.3791/51240
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发表时间：
2014-04-03
期刊：
影响因子：
--
通讯作者：
Melikian HE
中科院分区：
文献类型：
--
作者：
Gabriel LR;Wu S;Melikian HE
调节内吞运输是通过在分钟时间尺度上动态控制受体、离子通道和转运蛋白细胞表面呈现来促进各种神经调节事件的中心机制。控制单个蛋白质内吞运输的机制多种多样。研究贩运的分子基础主要依靠表面生物素化来定量测量膜蛋白表面表达响应外源刺激和基因操作的变化。然而，这种方法主要限于培养细胞，这可能无法忠实地反映成年神经元中发挥作用的生理相关机制。此外，培养细胞方法可能会低估贩运机制中特定区域的差异。在这里，我们描述了一种将细胞表面生物素化扩展到急性脑切片制备的方法。我们证明该方法提供了一种高保真方法来测量成体神经元膜蛋白表面水平的快速变化。这种方法可能在神经元内吞运输领域具有广泛的用途。
Regulated endocytic trafficking is the central mechanism facilitating a variety of neuromodulatory events, by dynamically controlling receptor, ion channel, and transporter cell surface presentation on a minutes time scale. There is a broad diversity of mechanisms that control endocytic trafficking of individual proteins. Studies investigating the molecular underpinnings of trafficking have primarily relied upon surface biotinylation to quantitatively measure changes in membrane protein surface expression in response to exogenous stimuli and gene manipulation. However, this approach has been mainly limited to cultured cells, which may not faithfully reflect the physiologically relevant mechanisms at play in adult neurons. Moreover, cultured cell approaches may underestimate region-specific differences in trafficking mechanisms. Here, we describe an approach that extends cell surface biotinylation to the acute brain slice preparation. We demonstrate that this method provides a high-fidelity approach to measure rapid changes in membrane protein surface levels in adult neurons. This approach is likely to have broad utility in the field of neuronal endocytic trafficking.
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影响因子：
25
作者：
Holton, KL;Loder, MK;Melikian, HE
通讯作者：
Melikian, HE
多巴胺能神经元中内体室和多巴胺转运蛋白的差异亚细胞分布。
影响因子：
3.5
作者：
Rao, Anjali;Simmons, Diana;Sorkin, Alexander
通讯作者：
Sorkin, Alexander
内吞作用未插入：多种进入细胞的方法。
影响因子：
44.1
作者：
Kumari S;Mg S;Mayor S
通讯作者：
Mayor S
影响因子：
5.3
作者：
Borgland, SL;Malenka, RC;Bonci, A
通讯作者：
Bonci, A
影响因子：
5.3
作者：
Sorkina, Tatiana;Miranda, Manuel;Sorkin, Alexander
通讯作者：
Sorkin, Alexander