Perturbation of regulated secretion in the pancreatic acinar cell line, AR42J.
Perturbation of regulated secretion in the pancreatic acinar cell line, AR42J.
复制标题
胰腺腺泡细胞系 AR42J 中调节分泌的扰动。
DOI：
10.1152/ajpgi.1992.262.2.g257
复制
发表时间：
1992
期刊：
影响因子：
--
通讯作者：
Jamieson,JD
中科院分区：
文献类型：
--
作者：
Sachs,E;Jamieson,JD
受调节的分泌途径包括响应激素刺激而加速蛋白质的释放、它们在分泌颗粒（SG）中的存在以及较长的细胞内停留时间。地塞米松诱导 AR42J 导致颗粒含量增加和对缩胆囊素 (CCK) 的反应性增加。我们使用[35S]蛋氨酸脉冲追踪方案研究了与将分泌蛋白分类到调节途径中有关的条件的影响，该方案检查了来自粗面内质网（RER）----SG，特别是来自高尔基复合体（GC）----SG的分泌蛋白的转运。后者在 20 摄氏度下进行追踪，使标记蛋白在反高尔基体中积累，然后转移到 37 摄氏度，在测试条件下启动它们向 SG 的转运。对 CCK-8 刺激的预存淀粉酶和在追踪过程中进入 SG 的新合成标记蛋白的释放进行定量，使我们能够检查扰动物对途径选定部分的影响。研究了酸性细胞内区室、细胞骨架、蛋白质合成、ATP 和温度对蛋白质在高尔基体进入调节途径之前和之后的影响。 NH4Cl、莫能菌素、叠氮化钠、20℃孵育和百日咳毒素可延迟RER----SG转运，但不影响淀粉酶排出。仅与 20 mM NH4Cl 或 1 microM 莫能菌素一起孵育才能抑制新合成蛋白质从晚期 GC----SG 的转移。 RER----高尔基体或高尔基体内运输似乎需要 ATP 和可能需要鸟苷 5'-三磷酸 (GTP) 结合蛋白。酸性区室似乎对于从 GC----SG 中分选分泌蛋白至关重要。
The regulated secretory pathway comprises accelerated discharge of proteins in response to hormonal stimuli, their presence in secretory granules (SG), and a long intracellular residence time. Dexamethasone induction of AR42J results in an increase in granule content and responsiveness to cholecystokinin (CCK). We studied the effects of conditions implicated in sorting of secretory proteins into the regulated pathway using [35S]methionine pulse-chase protocols that examine transport of secretory proteins from the rough endoplasmic reticulum (RER)----SG and specifically from the Golgi complex (GC)----SG. The latter uses a chase at 20 degrees C to allow accumulation of labeled proteins in the trans-Golgi, followed by a shift to 37 degrees C that initiates their transport to SG under test conditions. Quantitation of CCK-8-stimulated discharge of prestored amylase and of newly synthesized labeled proteins that have entered SG during the chase enables us to examine the effect of perturbants over selected parts of the pathway. The effects of acidic intracellular compartments, the cytoskeleton, protein synthesis, ATP, and temperature on pre- and post-Golgi entry of proteins into the regulated pathway were studied. NH4Cl, monensin, Na azide, incubation at 20 degrees C, and pertussis toxin retarded RER----SG transport without affecting amylase discharge. Only incubation with 20 mM NH4Cl or 1 microM monensin inhibited transfer of newly synthesized proteins from the late GC----SG. RER----Golgi or intra-Golgi transport thus appears to require ATP and possibly guanosine 5'-triphosphate (GTP)-binding proteins. Acidic compartments appear to be essential for sorting of secretory proteins from the GC----SG.