NEDD4-1 Regulates Migration and Invasion of Glioma Cells through CNrasGEF Ubiquitination In Vitro

Neuronal precursor cell-expressed developmentally down-regulated 4-1 (NEDD4-1) plays a great role in tumor cell growth, but its function and mechanism in cell invasive behavior are totally unknown. Here we report that NEDD4-1 regulates migration and invasion of malignant glioma cells via triggering ubiquitination of cyclic nucleotide Ras guanine nucleotide exchange factor (CNrasGEF) using cultured glioma cells. NEDD4-1 overexpression promoted cell migration and invasion, while its downregulation specifically inhibited them. However, NEDD4-1 did not affect the proliferation and apoptosis of glioma cells. NEDD4-1 physically interacted with CNrasGEF and promoted its poly-ubiquitination and degradation. Contrary to the effect of NEDD4-1, CNrasGEF downregulation promoted cell migration and invasion, while its overexpression inhibited them. Importantly, downregulation of CNrasGEF facilitated the effect of NEDD4-1-induced cell migration and invasion. Interestingly, aberrant up-regulated NEDD4-1 showed reverse correlation with CNrasGEF protein level but not with its mRNA level in glioma tissues. Combined with the in vitro results, the result of glioma tissues indicated post-translationally modification effect of NEDD4-1 on CNrasGEF. Our study suggests that NEDD4-1 regulates cell migration and invasion through ubiquitination of CNrasGEF in vitro.神经元前体细胞表达的发育下调蛋白4-1（Neuronal precursor cell-expressed developmentally down-regulated 4-1，NEDD4-1）在肿瘤细胞生长过程中发挥重要作用，但其在细胞侵袭行为中的功能与作用机制迄今仍完全不明。本研究通过培养的胶质瘤细胞证实，NEDD4-1可通过介导环状核苷酸Ras鸟苷酸交换因子（cyclic nucleotide Ras guanine nucleotide exchange factor，CNrasGEF）的泛素化，调控恶性胶质瘤细胞的迁移与侵袭能力。NEDD4-1过表达可促进细胞迁移与侵袭，而其表达下调则会特异性抑制这两种过程；但NEDD4-1并不会影响胶质瘤细胞的增殖与凋亡。NEDD4-1可与CNrasGEF发生直接物理相互作用，并促进其多泛素化修饰与降解。与NEDD4-1的作用相反，CNrasGEF表达下调可促进细胞迁移与侵袭，而过表达则会抑制该过程。值得注意的是，CNrasGEF表达下调可增强NEDD4-1诱导的细胞迁移与侵袭效应。有趣的是，在胶质瘤组织中，异常高表达的NEDD4-1与CNrasGEF的蛋白水平呈负相关，但与其mRNA水平无显著关联。结合体外实验结果，胶质瘤组织的相关实验结果证实了NEDD4-1对CNrasGEF的翻译后修饰调控作用。本研究表明，在体外环境中，NEDD4-1可通过泛素化调控CNrasGEF，进而影响细胞的迁移与侵袭能力。