Alix, a protein regulating endosomal trafficking, is involved in neuronal death
Alix, a protein regulating endosomal trafficking, is involved in neuronal death
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DOI：
10.1074/jbc.m309243200
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发表时间：
2004-01-16
影响因子：
4.8
通讯作者：
Sadoul, R
中科院分区：
文献类型：
--
作者：
Trioulier, Y;Torch, S;Sadoul, R
Alix/AIP1是一种细胞质蛋白，最初被认为是ALG-2的相互作用因子，ALG-2是细胞死亡所必需的钙结合蛋白。Alix最近也被定义为内溶酶体系统的调节因子。在这里，我们使用有丝分裂后的小脑神经元来测试Alix在caspase依赖性和非依赖性细胞死亡中的功能。事实上，这些神经元在25 mM含钾培养基中存活，但在细胞外钾降至5 mM后很快发生凋亡。与其他研究一致，我们表明，在K+剥夺后，半胱天冬酶被激活，但使用pancaspase抑制剂boc-天冬氨酸(OMe)-氟甲基酮抑制这些蛋白酶，对细胞存活没有影响。转染实验表明，Alix过表达足以诱导caspase激活，而其c端一半Alix- ct过表达可阻断caspase激活和K+剥夺后细胞死亡。我们还定义了结合ALG-2所需的c端富含脯氨酸结构域的12个氨基酸PXY重复序列。Alix或Alix- ct中该结构域的缺失消除了过表达蛋白对神经元存活的影响，表明alg -2结合区对Alix的死亡调节功能至关重要。总之，这些发现将Alix/ALG-2复合物定义为控制caspase依赖性和非依赖性途径的细胞死亡调节剂。他们还提出了内溶酶体系统和细胞死亡机制效应物之间的分子联系。
Alix/AIP1 is a cytoplasmic protein, which was first characterized as an interactor of ALG-2, a calcium-binding protein necessary for cell death. Alix has also recently been defined as a regulator of the endo-lysosomal system. Here we have used post-mitotic cerebellar neurons to test Alix function in caspase-dependent and - independent cell death. Indeed, these neurons survived when cultured in 25 mM potassium-containing medium but underwent apoptosis soon after the extracellular potassium was lowered to 5 mM. In agreement with other studies, we show that caspases are activated after K+ deprivation, but that inhibition of these proteases, using the pancaspase inhibitor boc-aspartyl(OMe)-fluoromethylketone, has no effect on cell survival. Transfection experiments demonstrated that Alix overexpression is sufficient to induce caspase activation, whereas overexpression of its C-terminal half, Alix-CT, blocks caspase activation and cell death after K+ deprivation. We also define a 12-amino acid PXY repeat of the C-terminal proline-rich domain necessary for binding ALG-2. Deletion of this domain in Alix or in Alix-CT abolished the effects of the overexpressed proteins on neuronal survival, demonstrating that the ALG-2-binding region is crucial for the death-modulating function of Alix. Overall, these findings define the Alix/ALG-2 complex as a regulator of cell death controlling both caspase-dependent and - independent pathways. They also suggest a molecular link between the endo-lysosomal system and the effectors of the cell death machinery.