Hansen H P, Dietrich S, Kisseleva T, Mokros T, Mentlein R, Lange H H, Murphy G, Lemke H
Departments of Biochemistry and Anatomy, University of Kiel, Kiel, Germany.
J Immunol. 2000 Dec 15;165(12):6703-9. doi: 10.4049/jimmunol.165.12.6703.
CD30 is a costimulatory receptor on activated lymphocytes and a number of human lymphoma cells. Specific ligation of membrane-bound CD30 or cellular stimulation by PMA results in a metalloproteinase-mediated down-regulation of CD30 and release of its soluble ectodomain (sCD30). In this report, it is demonstrated that PMA-induced CD30 cleavage from Karpas 299 cells was mediated by a membrane-anchored metalloproteinase which was active on intact cells following 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate extraction of membrane preparations. Moreover, CD30 shedding was blocked by the synthetic hydroxamic acid-based metalloproteinase inhibitor BB-2116 (IC(50), 230 nM) and the natural tissue inhibitor of metalloproteinases (TIMP)-3 (IC(50), 30 nM), but not by the matrix metalloproteinase inhibitors TIMP-1 and TIMP-2. This inhibition profile is similar to that of the TNF-alpha- converting enzyme (TACE) and, indeed, mRNA transcripts of the membrane-bound metalloproteinase-disintegrin TACE could be detected in Karpas 299 cells. The ectodomain of TACE was expressed in bacteria as a GST fusion protein (GST-TACE) which cleaved CD30 from the surface of Karpas 299 cells and concomitantly increased the level of sCD30 in the cell supernatants. Hence, TACE does not only control the release of TNF-alpha, but also that of sCD30.
CD30是活化淋巴细胞和许多人类淋巴瘤细胞上的共刺激受体。膜结合型CD30的特异性连接或佛波酯(PMA)对细胞的刺激会导致金属蛋白酶介导的CD30下调及其可溶性胞外域(sCD30)的释放。在本报告中,证明了PMA诱导的Karpas 299细胞中CD30的裂解是由一种膜锚定金属蛋白酶介导的,在用3-[(3-胆酰胺丙基)二甲基铵]-1-丙烷磺酸盐提取膜制剂后,该酶在完整细胞上具有活性。此外,合成的基于异羟肟酸的金属蛋白酶抑制剂BB-2116(IC50,230 nM)和天然金属蛋白酶组织抑制剂(TIMP)-3(IC50,30 nM)可阻断CD30的脱落,但基质金属蛋白酶抑制剂TIMP-1和TIMP-2则不能。这种抑制模式与肿瘤坏死因子-α转换酶(TACE)相似,实际上,在Karpas 299细胞中可检测到膜结合型金属蛋白酶-解整合素TACE的mRNA转录本。TACE的胞外域在细菌中作为GST融合蛋白(GST-TACE)表达,该蛋白可从Karpas 299细胞表面裂解CD30,并同时增加细胞上清液中sCD30的水平。因此,TACE不仅控制肿瘤坏死因子-α的释放,还控制sCD30的释放。