ANGI-14. A GSK-3/b-CATENIN/ARHGAP AXIS REGULATES GLIOBLASTOMA INVASION
ANGI-14. A GSK-3/b-CATENIN/ARHGAP AXIS REGULATES GLIOBLASTOMA INVASION
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ANGI-14。
DOI：
10.1093/neuonc/nox168.092
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发表时间：
2017
期刊：
影响因子：
15.9
通讯作者：
Brüning-Richardson A
中科院分区：
文献类型：
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作者：
Brüning-Richardson A
糖原合成酶激酶3 （GSK-3）已被认为是胶质母细胞瘤迁移和侵袭的潜在靶点。本文研究了GSK-3抑制对下游效应基因的影响，发现了一条新的GSK-3 /b-catenin/ARHGAP迁移轴，该轴驱动胶质母细胞瘤的侵袭。方法用GSK-3抑制剂6-溴靛红素-3′-肟（BIO）处理胶质母细胞瘤细胞进行表达阵列。通过2D和3D迁移分析、siRNA敲除和原位异种移植模型进行功能验证。研究了患者胶质母细胞瘤样本中的蛋白表达。结果微阵列数据分析发现，生物处理U251细胞后，GTPase调节因子ARHGAP基因家族成员的表达出现了异常。Western blotting和免疫染色在蛋白水平证实了这一点。siRNA处理表明ARHGAP29具有特异性的促迁移作用。暴露于BIO的U251细胞表现出迁移能力下降，表现为极性丧失和细胞速度降低。这与肌动蛋白丝的重排和b-连环蛋白从细胞表面转移到细胞核有关，在细胞核中，它被认为是GSK-3抑制的下游转录因子。然后我们发现，ICG001，一种b-catenin诱导转录的抑制剂，本质上模仿GSK-3的激活，诱导ARHGAP29的转录，并导致ARHGAP29蛋白表达的部分增加，b-catenin靶基因表达的降低和细胞迁移能力。体内研究以及来自胶质母细胞瘤患者的数据证实了ARHGAP29的促进迁移作用，证实了原位异种移植模型中侵袭前沿迁移细胞中蛋白水平升高及其与疾病复发的关联。我们首次证明了GSK-3激活和b-catenin核易位下游的信号事件，揭示了通过GTPase调节剂ARHGAP29的转录调节胶质母细胞瘤迁移的新途径。
INTRODUCTION Glycogen synthase kinase-3 (GSK-3) has been implicated as a potential target in glioblastoma migration and invasion. Here we investigated the effects of GSK-3 inhibition on downstream effector genes and discovered a novel GSK-3 /b-catenin/ARHGAP migration axis that drives glioblastoma invasion. METHODS Expression arrays of glioblastoma cells treated with the GSK-3 inhibitor 6-bromoindirubin-3’-oxime (BIO) were performed. Functional validation was carried out using 2D and 3D migration assays, siRNA knockdown and orthotopic xenograft models. Protein expression was investigated in patient glioblastoma samples. RESULTS Microarray data analysis identified deregulation of expression of members of the ARHGAP gene family of GTPase regulators after BIO treatment of U251 cells. This was confirmed at the protein level by Western blotting and immunostaining. siRNA treatment indicated a specific promigratory role for ARHGAP29. U251 cells exposed to BIO displayed a decrease in migration as evidenced by loss of polarity and reduction in cell velocity. This was associated with a rearrangement of actin filaments and b-catenin translocation from the cell surface to the nucleus, where it is known to act as a transcription factor downstream of GSK-3 inhibition. We then found that ICG001, an inhibitor of b-catenin-induced transcription, essentially mimicking GSK-3 activation, induced transcription of ARHGAP29 and led to a partial increase of ARHGAP29 protein expression, reduction of b-catenin target gene expression and the ability of cells to migrate. In vivo studies, as well as data from glioblastoma patients, confirmed a pro-migratory role for ARHGAP29 as evidenced by elevated protein levels in migratory cells at the invasive front in orthotopic xenograft models and its association with disease recurrence. DISCUSSION We demonstrate for the first time signalling events downstream of GSK-3 activation and b-catenin nuclear translocation which reveal a new pathway regulating glioblastoma migration through the transcription of the GTPase regulator ARHGAP29.