Actin filament disassembly is a sufficient final trigger for exocytosis in nonexcitable cells.
Actin filament disassembly is a sufficient final trigger for exocytosis in nonexcitable cells.
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DOI：
10.1083/jcb.128.4.589
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发表时间：
1995-02
期刊：
影响因子：
--
通讯作者：
Yin HL
中科院分区：
文献类型：
--
作者：
Muallem S;Kwiatkowska K;Xu X;Yin HL
虽然肌动蛋白细胞骨架与囊泡运输、对接和融合有关，但其作用位点以及与Ca（2+）介导的对接和融合机制激活的关系尚未阐明。在这项研究中，我们研究了肌动蛋白丝的作用，通过引入高度特异性的肌动蛋白单体结合蛋白，β-胸腺素或凝溶胶蛋白片段，到链球菌溶血素O-透性胰腺腺泡细胞调节胞吐。这些蛋白具有刺激和抑制作用。低浓度引起了快速和强大的胞吐与配置文件相比的初始阶段的调节胞吐，但没有提高[Ca 2 +]，甚至当[Ca 2 +]被钳位在低水平的EGTA。不需要额外的辅因子。肌动蛋白丝的直接可视化和定量显示，β-胸腺肽，像激动剂，诱导肌动蛋白解聚在顶膜胞吐发生。通过鬼笔环肽阻断肌动蛋白解聚或通过与外源性肌动蛋白复合来中和β-胸腺素可以防止胞吐作用。这些发现表明，皮层肌动蛋白网络作为一个显性负钳，阻止组成性胞吐。此外，肌动蛋白丝也有积极的作用。高浓度的肌动蛋白解聚蛋白抑制所有阶段的胞吐。抑制覆盖激动剂和所有下游效应测试的刺激，表明胞吐不能发生没有最小的肌动蛋白细胞骨架结构。
Although the actin cytoskeleton has been implicated in vesicle trafficking, docking and fusion, its site of action and relation to the Ca(2+)-mediated activation of the docking and fusion machinery have not been elucidated. In this study, we examined the role of actin filaments in regulated exocytosis by introducing highly specific actin monomer- binding proteins, the beta-thymosins or a gelsolin fragment, into streptolysin O-permeabilized pancreatic acinar cells. These proteins had stimulatory and inhibitory effects. Low concentrations elicited rapid and robust exocytosis with a profile comparable to the initial phase of regulated exocytosis, but without raising [Ca2+], and even when [Ca2+] was clamped at low levels by EGTA. No additional cofactors were required. Direct visualization and quantitation of actin filaments showed that beta-thymosin, like agonists, induced actin depolymerization at the apical membrane where exocytosis occurs. Blocking actin depolymerization by phalloidin or neutralizing beta- thymosin by complexing with exogenous actin prevented exocytosis. These findings show that the cortical actin network acts as a dominant negative clamp which blocks constitutive exocytosis. In addition, actin filaments also have a positive role. High concentrations of the actin depolymerizing proteins inhibited all phases of exocytosis. The inhibition overrides stimulation by agonists and all downstream effectors tested, suggesting that exocytosis cannot occur without a minimal actin cytoskeletal structure.