Stickle Natalie H, Chung Jacky, Klco Jeffery M, Hill Richard P, Kaelin William G, Ohh Michael
Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario M5S 1A8, Canada.
Mol Cell Biol. 2004 Apr;24(8):3251-61. doi: 10.1128/MCB.24.8.3251-3261.2004.
Functional inactivation of the von Hippel-Lindau (VHL) tumor suppressor gene is the cause of the familial VHL disease and most sporadic renal clear-cell carcinomas (RCC). pVHL has been shown to play a role in the destruction of hypoxia-inducible factor alpha (HIF-alpha) subunits via ubiquitin-mediated proteolysis and in the regulation of fibronectin matrix assembly. Although most disease-causing pVHL mutations hinder the regulation of the HIF pathway, every disease-causing pVHL mutant tested to date has failed to promote the assembly of the fibronectin matrix, underscoring its potential importance in VHL disease. Here, we report that a ubiquitin-like molecule called NEDD8 covalently modifies pVHL. A nonneddylateable pVHL mutant, while retaining its ability to ubiquitylate HIF, failed to bind to and promote the assembly of the fibronectin matrix. Expression of the neddylation-defective pVHL in RCC cells, while restoring the regulation of HIF, failed to promote the differentiated morphology in a three-dimensional growth assay and was insufficient to suppress the formation of tumors in SCID mice. These results suggest that NEDD8 modification of pVHL plays an important role in fibronectin matrix assembly and that in the absence of such regulation, an intact HIF pathway is insufficient to prevent VHL-associated tumorigenesis.
冯·希佩尔-林道（VHL）肿瘤抑制基因的功能失活是家族性VHL病和大多数散发性肾透明细胞癌（RCC）的病因。已证明pVHL通过泛素介导的蛋白水解作用参与缺氧诱导因子α（HIF-α）亚基的降解，并参与纤连蛋白基质组装的调节。尽管大多数致病的pVHL突变会阻碍HIF途径的调节，但迄今为止测试的每个致病pVHL突变体都未能促进纤连蛋白基质的组装，这突出了其在VHL病中的潜在重要性。在此，我们报告一种名为NEDD8的类泛素分子可共价修饰pVHL。一种不可被NEDD化的pVHL突变体，虽然保留了其泛素化HIF的能力，但未能结合并促进纤连蛋白基质的组装。在RCC细胞中表达缺乏NEDD化功能的pVHL，虽然恢复了对HIF的调节，但在三维生长试验中未能促进分化形态，并且不足以抑制SCID小鼠中的肿瘤形成。这些结果表明，pVHL的NEDD8修饰在纤连蛋白基质组装中起重要作用，并且在缺乏这种调节的情况下，完整的HIF途径不足以预防VHL相关的肿瘤发生。
