Sandsmark Danielle K, Zhang Huabiao, Hegedus Balazs, Pelletier Corey L, Weber Jason D, Gutmann David H
Department of Neurology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Cancer Res. 2007 May 15;67(10):4790-9. doi: 10.1158/0008-5472.CAN-06-4470.
Neurofibromatosis type 1 (NF1) is a common autosomal dominant tumor predisposition syndrome in which affected individuals develop astrocytic brain tumors (gliomas). To determine how the NF1 gene product (neurofibromin) regulates astrocyte growth and motility relevant to glioma formation, we have used Nf1-deficient primary murine astrocytes. Nf1(-/-) astrocytes exhibit increased protein translation and cell proliferation, which are mediated by Ras-dependent hyperactivation of the mammalian target of rapamycin (mTOR) protein, a serine/threonine protein kinase that regulates ribosomal biogenesis, protein translation, actin cytoskeleton dynamics, and cell proliferation. In this study, we show that Nf1-deficient astrocytes have fewer actin stress fibers and exhibit increased cell motility compared with wild-type astrocytes, which are rescued by pharmacologic and genetic mTOR inhibition. We further show that mTOR-dependent regulation of actin stress fiber formation, motility, and proliferation requires rapamycin-sensitive activation of the Rac1 GTPase but not elongation factor 4E-binding protein 1/S6 kinase. Nf1(-/-) astrocytes also exhibit increased protein translation and ribosomal biogenesis through increased expression of the nucleophosmin (NPM) nuclear-cytoplasmic shuttling protein. We found that NPM expression in Nf1(-/-) astrocytes was blocked by rapamycin in vitro and in vivo and that expression of a dominant-negative NPM mutant protein in Nf1(-/-) astrocytes rescued actin stress fiber formation and restored cell motility and proliferation to wild-type levels. Together, these data show that neurofibromin regulates actin cytoskeleton dynamics and cell proliferation through a mTOR/Rac1-dependent signaling pathway and identify NPM as a critical mTOR effector mediating these biological properties in Nf1-deficient astrocytes.
1型神经纤维瘤病（NF1）是一种常见的常染色体显性肿瘤易感综合征，患者会发生星形胶质细胞脑肿瘤（胶质瘤）。为了确定NF1基因产物（神经纤维瘤蛋白）如何调节与胶质瘤形成相关的星形胶质细胞生长和运动，我们使用了Nf1基因缺失的原代小鼠星形胶质细胞。Nf1（-/-）星形胶质细胞表现出蛋白质翻译增加和细胞增殖，这是由雷帕霉素哺乳动物靶蛋白（mTOR）的Ras依赖性过度激活介导的，mTOR是一种丝氨酸/苏氨酸蛋白激酶，可调节核糖体生物合成、蛋白质翻译、肌动蛋白细胞骨架动力学和细胞增殖。在本研究中，我们发现与野生型星形胶质细胞相比，Nf1基因缺失的星形胶质细胞的肌动蛋白应力纤维较少，细胞运动增加，而药物和基因mTOR抑制可挽救这种情况。我们进一步表明，mTOR对肌动蛋白应力纤维形成、运动和增殖的依赖性调节需要Rac1 GTP酶的雷帕霉素敏感激活，而不是真核翻译起始因子4E结合蛋白1/核糖体蛋白S6激酶（4E-BP1/S6K）。Nf1（-/-）星形胶质细胞还通过核磷蛋白（NPM）核质穿梭蛋白的表达增加，表现出蛋白质翻译和核糖体生物合成增加。我们发现，雷帕霉素在体外和体内均可阻断Nf1（-/-）星形胶质细胞中NPM的表达，并且在Nf1（-/-）星形胶质细胞中表达显性负性NPM突变蛋白可挽救肌动蛋白应力纤维的形成，并将细胞运动和增殖恢复到野生型水平。总之，这些数据表明神经纤维瘤蛋白通过mTOR/Rac1依赖性信号通路调节肌动蛋白细胞骨架动力学和细胞增殖，并确定NPM是在Nf1基因缺失的星形胶质细胞中介导这些生物学特性的关键mTOR效应器。
