SERIAL KILLING BY CYTOTOXIC T-LYMPHOCYTES - T-CELL RECEPTOR TRIGGERS DEGRANULATION, RE-FILLING OF THE LYTIC GRANULES AND SECRETION OF LYTIC PROTEINS VIA A NON-GRANULE PATHWAY
SERIAL KILLING BY CYTOTOXIC T-LYMPHOCYTES - T-CELL RECEPTOR TRIGGERS DEGRANULATION, RE-FILLING OF THE LYTIC GRANULES AND SECRETION OF LYTIC PROTEINS VIA A NON-GRANULE PATHWAY
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DOI：
10.1002/eji.1830250432
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发表时间：
1995-04-01
影响因子：
5.4
通讯作者：
GRIFFITHS, GM
中科院分区：
文献类型：
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作者：
ISAAZ, S;BAETZ, K;GRIFFITHS, GM
CD8(+)细胞毒性T淋巴细胞(CTL)克隆在同种异体靶细胞刺激下，开始合成裂解蛋白颗粒酶A、颗粒酶B和穿孔素。裂解蛋白储存在分泌颗粒中，在靶细胞识别后，T细胞受体(TCR)交联后释放出来。在裂解颗粒生物发生过程中，在同种异体CTL刺激后2-10天可检测到颗粒酶A的蛋白质合成。虽然颗粒酶A在此期间储存在裂解颗粒中，但合成的大部分颗粒酶A是直接从CTL分泌的。TCR触发的脱颗粒也会导致裂解蛋白的新合成，这可以被放线菌亚胺(CHX)抑制。一些新合成的裂解蛋白可以储存在细胞中，并重新填充颗粒。但是，颗粒酶A的活性和分泌的颗粒酶B的免疫印迹表明，多达三分之一的颗粒酶A和B可以通过构成分泌途径直接从CTL分泌，当三分之一的蛋白质无法获得颗粒靶向信号时，穿孔素也通过构成途径从自然杀伤细胞系YT和TCR交联后的CTL克隆中分泌。裂解蛋白的结构性分泌可被CHX和布雷菲尔丁A(BFA)阻断。虽然BFA不影响对识别靶点的定向杀伤，但它取消了旁观者杀伤，表明旁观者杀伤是由新合成的裂解蛋白通过非颗粒途径传递的。这些结果表明，在CTL杀伤多个靶点的同时，穿孔素/颗粒酶介导的裂解途径可以维持。我们发现，CTL不仅在杀伤过程中重新填充颗粒，而且还通过非颗粒介导的途径分泌裂解蛋白。
CD8(+) cytotoxic T lymphocyte (CTL) clones begin to synthesize the lytic proteins granzyme A, granzyme B and perforin after stimulation with allogeneic target cells. The lytic proteins are stored in the secretory granules which are released after cross-linking of the T cell receptor (TcR) upon target cell recognition. During lytic granule biogenesis granzyme A protein synthesis can be detected between 2 and 10 days after allogeneic stimulation of the CTL. Although granzyme A is stored in the lytic granules over this period, the majority of granzyme A synthesized is secreted directly from the CTL. TcR triggering of degranulation also results in new synthesis of the lytic proteins, which can be inhibited by cycloheximide (CHX). Some of the newly synthesized lytic proteins can be stored in the cell and refill the granules. But up to one third of granzymes A and B can be secreted directly from the CTL via the constitutive secretory pathway as shown by granzyme A enzymatic activity and immunoblots of secreted granzyme B, where one third of the protein fails to acquire the granule targeting signal, Perforin is also secreted via the constitutive pathway, both from the natural killer cell line, YT, and from CTL clones after TcR cross-linking. Constitutive secretion of the lytic proteins can be blocked by both CHX and brefeldin A (BFA). While BFA does not affect the directional killing of recognized targets, it abrogates bystander killing, indicating that bystander killing arises from newly synthesized lytic proteins delivered via a non-granule route.These results demonstrate that the perforin/granzyme-mediated lytic pathway can be maintained while CTL kill multiple targets. We show that CTL not only re-fill their granules during killing, but also secrete lytic proteins via a non-granule-mediated pathway.