Cortical filamentous actin disassembly and scinderin redistribution during chromaffin cell stimulation precede exocytosis, a phenomenon not exhibited by gelsolin.
Cortical filamentous actin disassembly and scinderin redistribution during chromaffin cell stimulation precede exocytosis, a phenomenon not exhibited by gelsolin.
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DOI：
10.1083/jcb.113.5.1057
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发表时间：
1991-06
期刊：
影响因子：
--
通讯作者：
Trifaró JM
中科院分区：
文献类型：
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作者：
Vitale ML;Rodríguez Del Castillo A;Tchakarov L;Trifaró JM
免疫荧光和细胞化学研究表明，丝状肌动蛋白主要定位于染色质细胞的皮质表面。有人认为，这些肌动蛋白丝网络作为分泌颗粒的屏障，阻碍了它们与质膜的接触。对嗜铬细胞的刺激产生肌动蛋白丝网络的分解，意味着屏障的移除。gelsolin和scinderin这两种依赖Ca（2+）的肌动蛋白断丝蛋白存在于染色质细胞的皮层表面，这表明细胞刺激可能会激活一种或多种肌动蛋白断丝蛋白，从而破坏肌动蛋白网络。因此，在培养的嗜铬细胞中，利用scinderin和gelsolin抗体以及丝状肌动蛋白探针罗丹明-phalloidin进行生化研究和荧光显微镜实验，研究scinderin、gelsolin和丝状肌动蛋白在细胞刺激过程中的分布情况，并将其可能的变化与儿茶酚胺的分泌联系起来。在这里，我们报告了在尼古丁刺激或K（+）诱发的去极化期间，皮层下的scinderin而不是gelsolin被重新分配，并且这种重新分配先于儿茶酚胺的分泌。烟碱受体介导烟碱素在斑块中的重排。细胞刺激会产生类似的纤维蛋白和丝状肌动蛋白分布模式。然而，在去除刺激后，scinderin皮层分布模式的恢复比F-actin重组要快，这表明scinderin在细胞皮层区域与F-actin以外的组分结合。我们还证明了外周肌动蛋白丝的分解和质下肌动蛋白的重新分布是钙依赖的事件。此外，针对多巴胺- β -羟化酶的抗体实验表明，胞吐位点优先定位于f -肌动蛋白分解区域。
Immunofluorescence and cytochemical studies have demonstrated that filamentous actin is mainly localized in the cortical surface of the chromaffin cell. It has been suggested that these actin filament networks act as a barrier to the secretory granules, impeding their contact with the plasma membrane. Stimulation of chromaffin cells produces a disassembly of actin filament networks, implying the removal of the barrier. The presence of gelsolin and scinderin, two Ca(2+)- dependent actin filament severing proteins, in the cortical surface of the chromaffin cells, suggests the possibility that cell stimulation brings about activation of one or more actin filament severing proteins with the consequent disruption of actin networks. Therefore, biochemical studies and fluorescence microscopy experiments with scinderin and gelsolin antibodies and rhodamine-phalloidin, a probe for filamentous actin, were performed in cultured chromaffin cells to study the distribution of scinderin, gelsolin, and filamentous actin during cell stimulation and to correlate the possible changes with catecholamine secretion. Here we report that during nicotinic stimulation or K(+)-evoked depolarization, subcortical scinderin but not gelsolin is redistributed and that this redistribution precedes catecholamine secretion. The rearrangement of scinderin in patches is mediated by nicotinic receptors. Cell stimulation produces similar patterns of distribution of scinderin and filamentous actin. However, after the removal of the stimulus, the recovery of scinderin cortical pattern of distribution is faster than F-actin reassembly, suggesting that scinderin is bound in the cortical region of the cell to a component other than F-actin. We also demonstrate that peripheral actin filament disassembly and subplasmalemmal scinderin redistribution are calcium-dependent events. Moreover, experiments with an antibody against dopamine-beta-hydroxylase suggest that exocytosis sites are preferentially localized to areas of F-actin disassembly.