Involvement of annexin II in exocytosis of lamellar bodies from alveolar epithelial type II cells
Involvement of annexin II in exocytosis of lamellar bodies from alveolar epithelial type II cells
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DOI：
10.1152/ajplung.1996.270.4.l668
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发表时间：
1996-04-01
影响因子：
4.9
通讯作者：
Zimmerman, UJP
中科院分区：
文献类型：
--
作者：
Liu, L;Wang, MS;Zimmerman, UJP
膜联蛋白是与胞吐作用有关的Ca 2+和磷脂结合蛋白家族。在本研究中，我们研究了选定的膜联蛋白参与板层体的胞吐作用，通过检查其脂质体的聚集特性和能力，从透化大鼠肺泡II型细胞重构表面活性剂分泌。膜联蛋白I，II，III和VI的证明在II型细胞的免疫印迹分析，但膜联蛋白IV和V没有发现。膜联蛋白I-IV介导的脂质体聚集在1 mM Ca 2+的存在下。然而，只有膜联蛋白II四聚体在10 μ M Ca 2+时具有聚集活性。膜联蛋白V和VI在任何Ca 2+浓度下（高达1 mM Ca 2+）具有可忽略的聚集活性。为了研究膜联蛋白的分泌重建，用40 μ M β-七叶皂苷透化分离的II型细胞。在这些条件下，透化的细胞向培养基中释放类似于30-40%的乳酸脱氢酶。在透化过程中，细胞膜联蛋白含量的一个未确定的部分丢失。然而，在透化的II型细胞中的板层体适当地用荧光染料尼罗红和奎纳克林染色，表明它们是完整的。这些透化的细胞分泌能力，因为磷脂酰胆碱（PC）分泌刺激0.2-1.0 μ M的Ca 2+。添加外源性膜联蛋白混合物增强PC分泌的透性IV型细胞在0.5 μ M的Ca 2+的最大刺激。六个纯化的膜联蛋白（I-VI）测试其能力，以重建分泌的透化细胞，只有膜联蛋白II是有效的。我们的研究结果表明，膜联蛋白II是必要的板层体的胞吐。
Annexins are a family of Ca2+- and phospholipid-binding proteins that have been implicated in exocytosis. In the present study, we investigated the participation of selected annexins in exocytosis of lamellar bodies by examining their liposome aggregation property and ability to reconstitute surfactant secretion from permeabilized rat lung alveolar type II cells. Annexins I, II, III, and VI were demonstrated in type II cells by immunoblot analysis, but annexin IV and V were not found. Annexins I-IV mediated liposome aggregation in the presence of 1 mM Ca2+. However, only annexin II tetramer had aggregation activity at 10 mu M Ca2+. Annexins V and VI had negligible aggregation activity at any Ca2+ concentrations (up to 1 mM Ca2+). To study reconstitution of secretion by annexins, isolated type II cells were permeabilized with 40 mu M beta-escin. Under these conditions, the permeabilized cells released similar to 30-40% lactic acid dehydrogenase into the medium. An undetermined fraction of cellular annexin content was lost during permeabilization. However, lamellar bodies in the permeabilized type II cells stained appropriately with the fluorescent dyes Nile red and quinacrine, indicating that they were intact. These permeabilized cells were secretion competent, since phosphatidylcholine (PC) secretion was stimulated by 0.2-1.0 mu M Ca2+. Addition of an exogenous annexin mixture enhanced PC secretion from the permeabilized type IV cells with maximal stimulation at 0.5 mu M Ca2+. Of six purified annexins (I-VI) tested for their ability to reconstitute secretion from the permeabilized cells, only annexin II was effective. Our results suggest that annexin II is necessary for exocytosis of lamellar bodies.