POSTTRANSLATIONAL REGULATION OF INSULIN-LIKE GROWTH-FACTOR BINDING PROTEIN-4 IN NORMAL AND TRANSFORMED HUMAN FIBROBLASTS - INSULIN-LIKE GROWTH-FACTOR DEPENDENCE AND BIOLOGICAL STUDIES
POSTTRANSLATIONAL REGULATION OF INSULIN-LIKE GROWTH-FACTOR BINDING PROTEIN-4 IN NORMAL AND TRANSFORMED HUMAN FIBROBLASTS - INSULIN-LIKE GROWTH-FACTOR DEPENDENCE AND BIOLOGICAL STUDIES
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DOI：
10.1172/jci116272
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发表时间：
1993-03-01
影响因子：
15.9
通讯作者：
ZAPF, J
中科院分区：
文献类型：
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作者：
CONOVER, CA;KIEFER, MC;ZAPF, J
胰岛素样生长因子结合蛋白-4（IGFBP-4）是一种分子量为24-26 kD的蛋白质，在体内和体外由多种细胞类型表达。用10 nM胰岛素样生长因子II（IGF-II）处理正常成人成纤维细胞24 h，通过条件培养基的Western配体印迹分析评估，内源性IGFBP-4减少85%。在无细胞条件下，将人成纤维细胞条件培养基（HFCM）与IGF-II孵育导致IGFBP-4类似的损失。这种IGFBP-4的后调节降低似乎是由于HFCM中的蛋白酶：（a）可以用特异性蛋白酶抑制剂或在4 ℃下孵育来防止;（B）重组人（rh）IGFBP-4的蛋白水解需要HFCM;（c）免疫印迹和放射性标记证实IGFBP-4裂解为18和14 kD IGFBP-4片段。 该蛋白酶对IGFBP-4具有特异性，并且严格依赖于IGFs的激活。IGF-II是测试的天然和突变IGF中最有效的，以0.25：1（IGF/IGFBP-4）的摩尔比诱导rhIGFBP 4的完全水解。猴病毒40转化的成人成纤维细胞也表达IGFBP 4和IGFBP-4蛋白酶，以及IGFBP 4蛋白水解的抑制剂。在生物学研究中，完整的rhIGFBP-4有效地抑制IGF-I刺激的[H-3]氨基异丁酸摄取，而蛋白水解的rhIGFBP-4没有抑制作用。总之，这些数据提供了一种新的IGF-依赖性IGFBP-4特异性蛋白酶，修改IGFBP-4的结构和功能的证据，并表明在其激活IGF-II的优先作用。IGFBP 4的翻译后调节可能为IGF-I和IGF-II协同控制局部细胞生长提供了一种手段。
Insulin-like growth factor binding protein-4 (IGFBP-4) is a 24-26-kD protein expressed by a variety of cell types in vivo and in vitro. Treatment of normal adult human fibroblasts with 10 nM insulin-like growth factor II (IGF-II) for 24 h resulted in an 85% decrease in endogenous IGFBP-4, as assessed by Western ligand blot analysis of the conditioned medium. Incubation of human fibroblast-conditioned medium (HFCM) with IGF-II under cell-free conditions led to a similar loss of IGFBP-4. This posttranslationally regulated decrease in IGFBP-4 appeared to be due to a protease in HFCM: (a) It could be prevented with specific protease inhibitors or incubation at 4-degrees-C; (b) proteolysis of recombinant human (rh) IGFBP-4 required HFCM; (c) immunoblotting and radiolabeling confirmed cleavage of IGFBP-4 into 18- and 14-kD IGFBP4 fragments. The protease was specific for IGFBP-4, and was strictly dependent on IGFs for activation. IGF-II was the most effective of the natural and mutant IGFs tested, inducing complete hydrolysis of rhIGFBP4 at a molar ratio of 0.25:1 (IGF/IGFBP-4). Simian virus 40-transformed adult human fibroblasts also expressed IGFBP4 and IGFBP-4 protease, as well as an inhibitor of IGFBP4 proteolysis. In biological studies, intact rhIGFBP4 potently inhibited IGF-I-stimulated [H-3]aminoisobutyric acid uptake, whereas proteolyzed rhIGFBP-4 had no inhibitory effect. In conclusion, these data provide evidence for a novel IGF-dependent IGFBP-4-specific protease that modifies IGFBP-4 structure and function, and indicate a preferential role for IGF-II in its activation. Posttranslational regulation of IGFBP4 may provide a means for cooperative control of local cell growth by IGF-I and IGF-II.