Inhibitory properties of cystatin F and its localization in U937 promonocyte cells
Inhibitory properties of cystatin F and its localization in U937 promonocyte cells
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DOI：
10.1111/j.1742-4658.2005.04594.x
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发表时间：
2005-03-01
期刊：
影响因子：
5.4
通讯作者：
Kos, J
中科院分区：
文献类型：
--
作者：
Langerholc, T;Zavasnik-Bergant, V;Kos, J
半胱氨酸蛋白酶抑制剂F是最近发现的几乎仅在免疫细胞中表达的II型半胱氨酸蛋白酶抑制剂。它存在于细胞内的溶酶体样囊泡，这表明在调节参与抗原呈递的木瓜蛋白酶样组织蛋白酶中的潜在作用。因此，在体外研究了半胱氨酸蛋白酶抑制剂F与其几种潜在靶标组织蛋白酶F、K、V、S、H、X和C的相互作用。半胱氨酸蛋白酶抑制剂F紧密地抑制组织蛋白酶F、K和V，Ki值在0.17 nm至0.35 nm范围内，而组织蛋白酶S和H以低100倍的亲和力被抑制（Ki约为30 nm）。外肽酶、组织蛋白酶C和X不受胱抑素F的抑制。为了研究抑制数据的生物学意义，通过共聚焦显微镜在U937前单核细胞中研究了半胱氨酸蛋白酶抑制剂F及其潜在靶点组织蛋白酶B、H、L、S、C和K的细胞内定位。虽然所有的酶都观察到囊泡染色，但只有组织蛋白酶H和X被发现与抑制剂共定位。这表明U937细胞中的胱抑素F可以作为组织蛋白酶H的蛋白水解活性的调节抑制剂，或者更可能的是，作为对组织蛋白酶的保护，所述组织蛋白酶被错误引导至含有内体/溶酶体囊泡的特定胱抑素F。半胱氨酸蛋白酶抑制剂F不与半胱氨酸蛋白酶抑制剂C共定位的发现表明这两种半胱氨酸蛋白酶抑制剂在U937细胞中具有不同的功能。
Cystatin F is a recently discovered type II cystatin expressed almost exclusively in immune cells. It is present intracellularly in lysosome-like vesicles, which suggests a potential role in regulating papain-like cathepsins involved in antigen presentation. Therefore, interactions of cystatin F with several of its potential targets, cathepsins F, K, V, S, H, X and C, were studied in vitro. Cystatin F tightly inhibited cathepsins F, K and V with K-i values ranging from 0.17 nm to 0.35 nm, whereas cathepsins S and H were inhibited with 100-fold lower affinities (K-i approximate to 30 nm). The exopeptidases, cathepsins C and X were not inhibited by cystatin F. In order to investigate the biological significance of the inhibition data, the intracellular localization of cystatin F and its potential targets, cathepsins B, H, L, S, C and K, were studied by confocal microscopy in U937 promonocyte cells. Although vesicular staining was observed for all the enzymes, only cathepsins H and X were found to be colocalized with the inhibitor. This suggests that cystatin F in U937 cells may function as a regulatory inhibitor of proteolytic activity of cathepsin H or, more likely, as a protection against cathepsins misdirected to specific cystatin F containing endosomal/lysosomal vesicles. The finding that cystatin F was not colocalized with cystatin C suggests distinct functions for these two cysteine protease inhibitors in U937 cells.