Upregulation of the neuron-specific K+/Cl- cotransporter expression by transcription factor early growth response 4
Upregulation of the neuron-specific K+/Cl- cotransporter expression by transcription factor early growth response 4
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DOI：
10.1523/jneurosci.4731-06.2006
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发表时间：
2006-12-27
影响因子：
5.3
通讯作者：
Airaksinen, Matti S.
中科院分区：
文献类型：
--
作者：
Uvarov, Pavel;Ludwig, Anastasia;Airaksinen, Matti S.
神经元特异性K（+）/Cl+协同转运蛋白（KCC 2）的表达仅限于CNS，并在神经元成熟过程中强烈上调，产生低的细胞内氯浓度，这是成年神经元快速突触抑制所需的。为了阐明KCC 2基因调控的机制，我们分析了KCC 2（别名Slc 12 a5）启动子和近端内含子1区域，并揭示了哺乳动物KCC 2基因中高度保守的10个候选转录因子结合位点。在这里，我们专注于这些因素之一，早期生长反应4（Egr 4），它显示了类似的发展上调中枢神经系统神经元KCC 2。在neuro-2a神经母细胞瘤细胞和培养的神经元中，Egr 4过表达强烈诱导含有Egr 4位点的KCC 2荧光素酶报告基因构建体（Egr 4（KCC 2））。点突变Egr 4 KCC 2显著降低Egr 4介导的诱导。插入Egr 4KCC 2到KCC 2基础启动子的内源性反向，但不是在相反的方向重建Egr 4介导的诱导。电泳迁移率变动分析证实了Egr 4与Egr 4KCC 2的特异性结合。干扰RNA介导的敲低Egr 4和Egr 4的显性负性亚型显著抑制培养神经元中KCC 2报告基因诱导和内源性KCC 2表达。总之，结果表明Egr 4在KCC 2基因表达的发育上调中起重要作用。
The expression of the neuron-specificK(+)/Cl+ cotransporter (KCC2) is restricted to the CNS and is strongly upregulated during neuronal maturation, yielding a low intracellular chloride concentration that is required for fast synaptic inhibition in adult neurons. To elucidate the mechanisms of KCC2 gene regulation, we analyzed the KCC2 (alias Slc12a5) promoter and proximal intron-1 regions and revealed 10 candidate transcription factor binding sites that are highly conserved in mammalian KCC2 genes. Here we focus on one of these factors, early growth response 4 (Egr4), which shows a similar developmental upregulation in CNS neurons as KCC2. KCC2 luciferase reporter constructs containing the Egr4 site (Egr4(KCC2)) were strongly induced by Egr4 overexpression in neuro-2a neuroblastoma cells and in cultured neurons. Egr4-mediated induction was decreased significantly by point-mutating the Egr4KCC2. Insertion of Egr4KCC2 into the KCC2 basal promoter in the endogenous reverse, but not in the opposite, orientation reestablished Egr4-mediated induction. Electrophoretic mobility shift assay confirmed specific Egr4 binding to Egr4KCC2. Interference RNA-mediated knock-down of Egr4 and a dominant-negative isoform of Egr4 significantly inhibited KCC2 reporter induction and endogenous KCC2 expression in cultured neurons. Together, the results indicate an important role for Egr4 in the developmental upregulation of KCC2 gene expression.