Receptor-mediated restoration of exocytotic capacity

我们证明了嗜碱性白血病（RBL-2H3）细胞在接收到适当的信号之前具有阻止胞外分泌的机制，并且细胞骨架，特别是肌动蛋白网络的意义在于其阻止胞外分泌。在制备rbr -m3细胞的过程中，我们也证明了Rho在高亲和力IgE受体（FcepsilonRI）和m3毒蕈碱乙酰胆碱受体（mAChRs）等多种受体介导的胞外分泌中起重要作用，并表明Rho通过调节肌动蛋白重组作为胞外分泌的整合点。这种受体介导的肌动蛋白网络重组，一个独立于Ca^<2+>的过程，可能揭示了诱导胞吐的能力。用编码人m3 machr的cDNA转染RBL-2H3细胞，研究其分泌的脱敏性。RBL-m3细胞在无Ca^<2+>的培养基中暴露于100 muM carbachol 30 min，可抑制随后添加carbachol和交联FcepsilonRI诱导的…More d分泌。RBL-m3细胞脱敏没有任何m3 mAChR修饰或受体与磷脂酶c之间的功能解偶联。我们的新发现是RBL-m3细胞的异源脱敏发生在细胞内钙浓度升高的远端。在本研究中，RBL-m3细胞的脱敏处理是在没有细胞外Ca^<2+>的情况下进行的，这表明异源和同源的分泌物脱敏是由于持续刺激肌动蛋白相关的“启动”信号而发生的。RBL-m3细胞在不含Ca^<2+>的培养基中加入10 μ m的carbachol后，出现了膜皱褶，而随后加入carbachol后，脱敏细胞没有出现这种皱褶。这些发现表明，碳水化合物诱导的异源和同源分泌物脱敏参与了导致膜褶皱的信号通路的负调控。综上所述，激动剂激活的肌动蛋白重组的上下调节可能是胞外反应能力受到调节的一种方式。少

We demonstrated that basophilic leukemia (RBL-2H3) cells possess a mechanism for preventing exocytosis until an appropriate signal is received by the cell, and that the significance of the cytoskeleton, particularly the actin network, lies in its preventing exocytosis. We also demonstrated in the preparation of RBL-m3 cells that Rho plays an essential role in the exocytosis mediated by a multiple type of receptor including high affinity IgE receptors (FcepsilonRI) and m3 muscarinic acetylcholine receptors (mAChRs) and suggested that Rho functions as an integration point for exocytosis by regulating actin reorganization. Such a receptor-mediated reorganization of the actin network, a process independent of Ca^<2+>, may reveal the capacity to induce exocytosis.The desensitization of secretion was investigated by transfecting RBL-2H3 cells with cDNA encoding the human m3 mAChRs. Exposure of RBL-m3 cells for 30 min to 100 muM carbachol in Ca^<2+>-free medium inhibited both secretion induce … More d by subsequent addition of carbachol and by cross-linking FcepsilonRI.RBL-m3 cells were desensitized without any modification of m3 mAChR or functional uncoupling between the receptor and phospholipase C.Our novel finding was that the heterologous desensitization of RBL-m3 cells occurred at the steps distal to the rise in concentration of intracellular calcium. Desensitizing treatment of RBL-m3 cells in this study was performed in the absence of extracellular Ca^<2+>, indicating that heterologous as well as homologous desensitization of secretion occurred as a consequence of persistent stimulation of the actin-related 'priming' signaling as described earlier. Incubation of RBL-m3 cells with 10 muM carbachol in Ca^<2+>-free medium developed membrane ruffling, while desensitizad cells failed to develop such ruffling with the subsequent addition of carbachol. These findings indicate that carbachol-induced heterologous as well as homologous desensitization of secretion involves negative regulation of the signaling pathway leading to membrane ruffling.In conclusion, it is likely that up- and down-regulation of agonist-activated actin reorganization leading to membrane ruffling is one way in which the capacity of exocytotic responses is regulated. Less