Structural requirements for the growth factor activity of the amino-terminal domain of urokinase.
Structural requirements for the growth factor activity of the amino-terminal domain of urokinase.
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DOI：
10.1016/s0021-9258(19)49691-9
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发表时间：
1992-07
期刊：
影响因子：
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通讯作者：
S. Rabbani;A. Mazar;S. Bernier;M. Haq;I. Bolivar;J. Henkin;David Goltzman
中科院分区：
文献类型：
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作者：
S. Rabbani;A. Mazar;S. Bernier;M. Haq;I. Bolivar;J. Henkin;David Goltzman
检测了蛋白水解活性失活的高分子量尿激酶型纤溶酶原激活剂（uPA）（二异丙基氟磷酸（DFP）-uPA）、其氨基末端片段（ATF，氨基酸（aa）1-143）以及岩藻糖基化和去岩藻糖基化生长因子结构域（GFD，aa 4-43）在人SaOS-2骨肉瘤细胞和U-937淋巴瘤细胞中的促生长作用和结合。DFP-uPA、ATF以及岩藻糖基化和去岩藻糖基化的GFD都能够与125 I-ATF竞争结合SaOS-2和U-937细胞。DFP-uPA、ATF和岩藻糖基化GFD在SaOS-2细胞中也具有促有丝分裂作用并增加细胞数量。然而，去岩藻糖基化GFD在SaOS-2细胞中是非促有丝分裂的，并且不刺激细胞增殖，即使它以与岩藻糖基化GFD相当的方式结合到这些细胞上。从大肠杆菌中表达并纯化的非糖基化高分子量uPA抑制125 I-ATF与SaOS-2细胞的结合，但也是非促有丝分裂的。在用能够在SaOS-2细胞中引发促有丝分裂反应的uPA形式处理的U-937细胞中未观察到促有丝分裂活性。蛋白水解制备的Kringle结构域（aa 47-135）和低分子量uPA（aa 144-411）不会竞争125 I-ATF结合，并且在所测试的两种细胞系中均不会引起任何促有丝分裂反应。此外，组织纤溶酶原激活剂（tPA），已被证明是同源的uPA在其生长因子域，也是岩藻糖基化，不抑制125 I-ATF结合，也不引起任何促有丝分裂反应。这些结果表明，涉及与uPA受体结合的GFD也负责SaOS-2细胞中的生长因子样活性，并且该结构域内Thr 18处的岩藻糖基化可以作为引发该应答的分子触发剂。
High molecular weight urokinase-type plasminogen activator (uPA) in which proteolytic activity was inactivated (diisopropyl fluorophosphate (DFP)-uPA), its amino-terminal fragment (ATF, amino acids (aa) 1-143), and fucosylated and defucosylated growth factor domains (GFD, aa 4-43) were tested for growth-promoting effects and binding in human SaOS-2 osteosarcoma cells and U-937 lymphoma cells. DFP-uPA, ATF, and both the fucosylated and defucosylated GFD were capable of competing with 125I-ATF for binding to both SaOS-2 and U-937 cells. DFP-uPA, ATF, and fucosylated GFD were also mitogenic in SaOS-2 cells and increased cell numbers. However, defucosylated GFD was nonmitogenic in SaOS-2 cells and did not stimulate cell proliferation, even though it bound to these cells in a manner equivalent to the fucosylated GFD. A nonglycosylated high molecular weight uPA expressed and purified from Escherichia coli inhibited 125I-ATF binding to SaOS-2 cells but was also nonmitogenic. No mitogenic activity was observed in U-937 cells treated with the uPA forms capable of eliciting a mitogenic response in SaOS-2 cells. Proteolytically prepared kringle domain (aa 47-135) and low molecular weight uPA (aa 144-411) did not compete for 125I-ATF binding and did not elicit any mitogenic response in either of the cell lines tested. In addition, tissue plasminogen activator (tPA), which has been shown to be homologous to uPA in its growth factor domain and is also fucosylated, did not inhibit 125I-ATF binding nor elicit any mitogenic response. These results demonstrate that the GFD, implicated in binding to the uPA receptor, is also responsible for growth factor like activity in SaOS-2 cells and that the fucosylation at Thr18 within this domain may serve as a molecular trigger in eliciting this response.