Phospholipases D1 and D2 regulate different phases of exocytosis in mast cells
Phospholipases D1 and D2 regulate different phases of exocytosis in mast cells
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DOI：
10.4049/jimmunol.168.11.5682
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发表时间：
2002-06-01
影响因子：
4.4
通讯作者：
Beaven, MA
中科院分区：
文献类型：
--
作者：
Choi, WS;Kim, YM;Beaven, MA
大鼠肥大细胞系RBL-2H3含有磷脂酶D（PLD）1和PLD 2。以前对该细胞系的研究表明，表达的PLD 1和PLD 2都被分泌刺激物强烈激活。我们现在显示，通过使用标记有增强型绿色荧光蛋白的PLD，PLD 1，这在很大程度上是与分泌颗粒，重新分配到质膜在受刺激的细胞中的过程，让人想起胞吐和融合的颗粒与质膜。这些过程和颗粒的分泌被PLD 1的催化失活突变体的表达或50 mM 1-丁醇而不是叔丁醇的存在所抑制，这表明这些事件依赖于PLD 1的催化活性。值得注意的是，霍乱毒素诱导PLD 1标记的颗粒易位到质膜，但不与质膜或分泌物的颗粒融合。随后用Ag或毒胡萝卜素刺激钙内流导致PLD 1快速重新分布到质膜并加速分泌。同样值得注意的是，PLD 1在刺激后4小时内从质膜再循环回到颗粒。相反，PLD 2主要局限于质膜，但它也参与分泌过程，因为催化失活的PLD 2的表达也阻断分泌。这些数据表明，一个两步的过程：易位的颗粒细胞周边，调节颗粒相关的PLD 1，和钙依赖性融合的颗粒与质膜，调节质膜相关的PLD 2和可能的PLD 1。
The rat mast cell line RBL-2H3 contains both phospholipase D (PLD)1 and PLD2. Previous studies with this cell line indicated that expressed PLD1 and PLD2 are both strongly activated by stimulants of secretion. We now show by use of PLDs tagged with enhanced green fluorescent protein that PLD1, which is largely associated with secretory granules, redistributes to the plasma membrane in stimulated cells by processes reminiscent of exocytosis and fusion of granules with the plasma membrane. These processes and secretion of granules are suppressed by expression of a catalytically inactive mutant of PLD1 or by the presence of 50 mM 1-butanol but not tent-butanol, an indication that these events are dependent on the catalytic activity of PLD1. Of note, cholera toxin induces translocation of PLD1-labeled granules to the plasma membrane but not fusion of granules with plasma membrane or secretion. Subsequent stimulation of calcium influx with Ag or thapsigargin leads to rapid redistribution of PLD1 to the plasma membrane and accelerated secretion. Also of note, PLD1 is recycled from plasma membrane back to granules within 4 h of stimulation. PLD2, in contrast, is largely confined to the plasma membrane, but it too participates in the secretory process, because expression of catalytically inactive PLD2 also blocks secretion. These data indicate a two-step process: translocation of granules to the cell periphery, regulated by granule-associated PLD1, and a calcium-dependent fusion of granules with the plasma membrane, regulated by plasma membrane-associated PLD2 and possibly PLD1.