EXTRACELLULAR RELEASE OF LYMPHOCYTE CYTOLYTIC PORE-FORMING PROTEIN (PERFORIN) AFTER IONOPHORE STIMULATION
EXTRACELLULAR RELEASE OF LYMPHOCYTE CYTOLYTIC PORE-FORMING PROTEIN (PERFORIN) AFTER IONOPHORE STIMULATION
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DOI：
10.1073/pnas.83.15.5668
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发表时间：
1986-08-01
影响因子：
11.1
通讯作者：
COHN, ZA
中科院分区：
文献类型：
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作者：
YOUNG, JDE;LEONG, LG;COHN, ZA
淋巴细胞颗粒的细胞溶孔形成蛋白（PFP，穿孔素）最近被分离和表征。分离的颗粒和纯化的PFP表达的裂解活性都需要Ca2+的存在。在这里，我们报道了Ca2+离子载体A23187刺激淋巴细胞后PFP的细胞外释放，该细胞脱颗粒。分泌的蛋白质与脂质结合形成结构和功能通道和超分子复合物，部分抵抗十二烷基硫酸钠和还原剂的解离。释放物质的免疫印迹显示与小鼠PFP和人补体成分C9特异性抗体阳性，表明这两个分子之间存在交叉反应性。利用这些特异性抗体作为免疫吸附剂，从受刺激细胞的上清液中亲和纯化淋巴细胞PFP。PFP的细胞外释放与在0.15 M NaCl中同时形成电导为550- 600ps的功能性离子非选择通道有关。在没有细胞外Ca2+的情况下，观察到最大释放活性的15%。Ca2+似乎需要引起淋巴细胞的分泌和释放的PFP组装成管状聚合物。在淋巴细胞杀伤细胞的过程中，PFP也可能产生类似的分泌，导致其聚集在靶膜上形成管状跨膜病变。
The cytolytic pore-forming protein (PFP, perforin) of lymphocyte granules has recently been isolated and characterized. The lytic activities expressed by both the isolated granules and the purified PFP require the presence of Ca2+. Here, we report on the extracellular release of PFP after stimulation of lymphocytes with the Ca2+ ionophore A23187, which degranulates the cells. The secreted protein associates with lipid to form structural and functional channels and supramolecular complexes that partially resist dissociation by sodium dodecyl sulfate and reducing agents. Immunoblots of the released material reveal positive identification with antibodies specific for mouse PFP and human complement component C9, indicating cross-reactivity between these two molecules. By using these specific antibodies as immunoadsorbents, the lymphocyte PFP has been affinity purified from the supernatant of stimulated cells. The extracellular release of PFP is associated with simultaneous formation of functional ion-nonselective channels with conductances of 550-600 pS in 0.15 M NaCl, as measured in planar model bilayers. In the absence of extracellular Ca2+, 15% of the maximal release activity is observed. Ca2+ appears to be required to elicit both secretion by lymphocytes and the assembly of the released PFP into tubular polymers. Similar secretion of PFP may occur during cell killing by lymphocytes, resulting in its assembly on target membranes to form tubular transmembrane lesions.