G(i) regulation of secretory vesicle swelling examined by atomic force microscopy
G(i) regulation of secretory vesicle swelling examined by atomic force microscopy
复制标题
DOI：
10.1073/pnas.94.24.13317
复制
发表时间：
1997-11-25
影响因子：
11.1
通讯作者：
Sritharan, KC
中科院分区：
文献类型：
--
作者：
Jena, BP;Schneider, SW;Sritharan, KC
在过去的十年中，已经发现几种单体和异三聚体鸟嘌呤核苷酸结合蛋白与分泌囊泡相关并参与胞吐作用，囊泡体积也被认为在分泌囊泡在质膜上的融合中发挥调节作用，然而，鸟嘌呤核苷酸结合蛋白功能的分子机制和胞吐过程中分泌囊泡体积的调节在这项研究中，我们报告了分泌囊泡膜与异源三聚体GTP结合蛋白G的α亚基的关系。（α i3），并暗示其参与囊泡肿胀，使用原子力显微镜结合共聚焦显微镜，我们能够研究分离的酶原颗粒的动力学，外分泌胰腺中的分泌囊泡，将酶原颗粒暴露于GTP导致囊泡高度增加15-25%，如通过原子力显微镜测量的，并且囊泡直径增加类似，如通过共聚焦显微镜测定的，Mas 7，一种已知刺激G（i）蛋白的活性mastoparan类似物，在GTP、NaF和Mas 7存在下，囊泡大小的增加是不可逆的，并且是KCl敏感的，Ca 2+对酶原颗粒大小没有影响。结果表明，定位于分泌囊泡膜中的G（α 13）蛋白介导囊泡肿胀，这是囊泡在细胞质膜融合的潜在重要先决条件。
In the last decade, several monomeric and heterotrimeric guanine nucleotide binding proteins have been identified to associate with secretory vesicles and to be implicated in exocytosis, Vesicle volume also has been proposed to play a regulatory role in secretory vesicle fusion at the plasma membrane, However, the molecular mechanism of function of the guanine nucleotide binding proteins and of the regulation of secretory vesicle volume in the exocytotic process remains unclear, In this study, we report association of the secretory vesicle membrane with the alpha subunit of a heterotrimeric GTP binding protein G(alpha i3) and implicate its involvement in vesicle swelling, Using an atomic force microscope in combination with confocal microscopy, we were able to study the dynamics of isolated zymogen granules, the secretory vesicles in exocrine pancreas, Exposure of zymogen granules to GTP resulted in a 15-25% increase in vesicle height as measured by the atomic force microscope and a similar increase in vesicle diameter as determined by confocal microscopy, Mas7, an active mastoparan analog known to stimulate G(i) proteins, was found to stimulate the GTPase activity of isolated zymogen granules and cause swelling, Increase in vesicle size in the presence of GTP, NaF, and Mas7 were irreversible and KCl-sensitive, Ca2+ had no effect on zymogen granule size, Taken together, the results indicate that G(alpha i3) protein localized in the secretory vesicle membrane mediates vesicle swelling, a potentially important prerequisite for vesicle fusion at the cell plasma membrane.