Rapid phosphorylation of histone H2A.X following ionotropic glutamate receptor activation
Rapid phosphorylation of histone H2A.X following ionotropic glutamate receptor activation
复制标题
DOI：
10.1111/j.1460-9568.2006.04768.x
复制
发表时间：
2006-05-01
影响因子：
3.4
通讯作者：
Kondratyev, Alexei
中科院分区：
文献类型：
--
作者：
Crowe, Samantha L.;Movsesyan, Vilen A.;Kondratyev, Alexei
离子型谷氨酸受体的过度激活增加氧化应激，导致神经损伤（如缺血和癫痫发作）后观察到的神经元死亡。翻译后组蛋白修饰可能是检测和修复氧化应激（包括DNA损伤）引起的损伤的关键介质，因此可能影响以过量谷氨酸释放为特征的损伤后神经元的存活。在非神经元细胞中，组蛋白变体H2A.X（称为γ-H2 AX）的磷酸化在DNA双链断裂后迅速发生。我们研究了γ-H2 AX形成大鼠皮层神经元（天在体外14）激活后的N-甲基-D-天冬氨酸（NMDA）或α-氨基-3-羟基-5-甲基-4-异恶唑丙酸（AMPA）/红藻氨酸谷氨酸受体，使用荧光免疫组织化学技术。此外，我们评估了γ-H2 AX“病灶”与双链断裂修复蛋白Mre 11的共定位，以提供该DNA损伤反应途径激活的进一步证据。在这里，我们表明，离子型谷氨酸受体的最低限度的细胞毒性刺激足以引起神经元中的γ-H2 AX，和NMDA诱导的γ-H2 AX灶的形成衰减预处理的抗氧化剂，维生素E，和细胞内钙螯合剂，BAPTA-AM。此外，γ-H2 AX灶的子集与Mre 11共定位，表明至少一部分γ-H2 AX灶是损伤依赖性的。谷氨酸受体激活后γ-H2 AX诱导的程度对应于我们在常规DNA损伤剂[即非致死剂量的γ-辐射（1戈伊）和过氧化氢（10 μ M）]后观察到的增加。这些数据表明，不一定导致神经元死亡的损伤诱导DNA损伤诱发的染色质修饰，γ-H2 AX，并暗示组蛋白改变在确定神经损伤后神经元的脆弱性的作用。
Excessive activation of ionotropic glutamate receptors increases oxidative stress, contributing to the neuronal death observed following neurological insults such as ischemia and seizures. Post-translational histone modifications may be key mediators in the detection and repair of damage resulting from oxidative stress, including DNA damage, and may thus affect neuronal survival in the aftermath of insults characterized by excessive glutamate release. In non-neuronal cells, phosphorylation of histone variant H2A.X (termed gamma-H2AX) occurs rapidly following DNA double-strand breaks. We investigated gamma-H2AX formation in rat cortical neurons (days in vitro 14) following activation of N-methyl-D-aspartate (NMDA) or alpha-amino-3-hydroxyl-5-methyl-4-isoxazolepropionic acid (AMPA)/kainate glutamate receptors using fluorescent immunohistochemical techniques. Moreover, we evaluated the co-localization of gamma-H2AX 'foci' with Mre11, a double-strand break repair protein, to provide further evidence for the activation of this DNA damage response pathway. Here we show that minimally cytotoxic stimulation of ionotropic glutamate receptors was sufficient to evoke gamma-H2AX in neurons, and that NMDA-induced gamma-H2AX foci formation was attenuated by pretreatment with the antioxidant, Vitamin E, and the intracellular calcium chelator, BAPTA-AM. Moreover, a subset of gamma-H2AX foci co-localized with Mre11, indicating that at least a portion of gamma-H2AX foci is damage dependent. The extent of gamma-H2AX induction following glutamate receptor activation corresponded to the increases we observed following conventional DNA damaging agents [i.e. non-lethal doses of gamma-radiation (1 Gy) and hydrogen peroxide (10 mu M)]. These data suggest that insults not necessarily resulting in neuronal death induce the DNA damage-evoked chromatin modification, gamma-H2AX, and implicate a role for histone alterations in determining neuronal vulnerability following neurological insults.