Puromycin-insensitive leucyl-specific aminopeptidase (PILSAP) binds and catalyzes PDK1, allowing VEGF-stimulated activation of S6K for endothelial cell proliferation and angiogenesis
Puromycin-insensitive leucyl-specific aminopeptidase (PILSAP) binds and catalyzes PDK1, allowing VEGF-stimulated activation of S6K for endothelial cell proliferation and angiogenesis
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DOI：
10.1182/blood-2003-12-4260
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发表时间：
2004-10-15
期刊：
影响因子：
20.3
通讯作者：
Sato, Y
中科院分区：
文献类型：
--
作者：
Yamazaki, T;Akada, T;Sato, Y
purromycin -insensitive leucyl-specific aminopeptidase （PILSAP）通过调节内皮细胞（ECs）的增殖和迁移在血管生成中起重要作用。在这里，我们描述了PILSAP调节血管内皮生长因子（VEGF）刺激的内皮细胞增殖的机制。特异性消除PILSAP表达或其酶活性可抑制内皮细胞中vegf刺激的G1/S转变。G1阻滞与细胞周期蛋白依赖性激酶4/6 （CDK4/6）活性降低和视网膜母细胞瘤（Rb）蛋白磷酸化相关。对CDK4/6上游信号分子的分析显示，S6激酶（S6K）的激活受到PILSAP的影响，而磷脂酰肌醇-3激酶（PI3K）、Akt和细胞外信号相关激酶1/2 （ERK1/2）的激活则不受影响。我们进一步证明，PILSAP结合磷脂酰肌醇依赖性激酶1 (PDK1)，并从其n端去除9个氨基酸，这使得S6K在VEGF刺激下与PDK1和PILSAP结合。我们构建了缺乏氨基肽酶活性但结合PDK1的突变体PILSAP。突变的PILSAP以显性阴性的方式消除了S6K在VEGF刺激下的激活。PDK1的n端截断形式消除了突变体PILSAP的显性负作用。最后，在ECs中引入突变的PILSAP基因可以抑制血管生成并延缓肿瘤生长。这些结果表明，PILSAP通过PDK1的结合和修饰，在内皮细胞的细胞周期进程和血管生成中起着至关重要的作用。(C) 2004年由美国血液病学会出版。
Puromycin-insensitive leucyl-specific aminopeptidase (PILSAP) plays an important role in angiogenesis by regulating the proliferation and migration of endothelial cells (ECs). Here we characterize the mechanism by which PILSAP regulates the vascular endothelial growth factor (VEGF)-stimulated proliferation of ECs. The specific elimination of PILSAP expression or its enzymatic activity inhibited VEGF-stimulated G1/S transition in ECs. This G1 arrest correlated with reduced cyclin dependent kinase 4/6 (CDK4/6) activity and retinoblastoma (Rb) protein phosphorylation. Analyses of signaling molecules upstream of CDK4/6 revealed that S6 kinase (S6K) activation was affected by PILSAP, whereas that of phosphatidylinositol-3 kinase (PI3K), Akt, and extracellular signal-related kinase 1/2 (ERK1/2) was not. We further demonstrated that PILSAP bound phosphatidylinositol-dependent kinase 1 (PDK1) and removed 9 amino acids from its N-terminus, which allowed S6K to associate with PDK1 and PILSAP upon VEGF stimulation. We constructed mutant PILSAP, which lacked the aminopeptidase activity but bound PDK1. Mutant PILSAP abrogated S6K activation upon VEGF stimulation in a dominant-negative manner. An N-terminal truncated form of PDK1 abolished the dominant-negative effect of mutant PILSAP. Finally, the introduction of a mutated PILSAP gene in ECs inhibited angiogenesis and retarded tumor growth in vivo. These results indicate that PILSAP plays a crucial role in the cell cycle progression of ECs and angiogenesis via the binding and modification of PDK1. (C) 2004 by The American Society of Hematology.