Identification of T-cadherin as a novel target of DNA methyltransferase 3B and its role in the suppression of nerve growth factor-mediated neurite outgrowth in PC12 cells (Retracted article. See vol. 293, pg. 3592, 2018)
Identification of T-cadherin as a novel target of DNA methyltransferase 3B and its role in the suppression of nerve growth factor-mediated neurite outgrowth in PC12 cells (Retracted article. See vol. 293, pg. 3592, 2018)
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DOI：
10.1074/jbc.m513278200
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发表时间：
2006-05-12
影响因子：
4.8
通讯作者：
Jacob, ST
中科院分区：
文献类型：
--
作者：
Bai, SM;Ghoshal, K;Jacob, ST
以前，我们表明，DNA甲基转移酶3b（Dnmt 3b）是所需的神经生长因子（NGF）诱导分化的PC 12细胞的神经元表型。本研究通过染色质免疫沉淀法（ChIP）鉴定了T-cadherin（T-Cad）为Dnmt 3b的靶点之一。结合亚硫酸氢盐限制性酶切分析和亚硫酸氢盐测序结果表明，T-Cad启动子在PC 12细胞中有稀疏的甲基化。ChIP-CHOP分析表明，Dnmt 3b与T-Cad启动子相关，而与其甲基化状态无关。反义或小干扰RNA去除Dnmt 3b后，T-Cad的mRNA和蛋白水平均显著升高。Dnmt 3b对T-Cad启动子活性的抑制不依赖于其催化活性，这与Dnmt 3b耗尽的细胞中T-Cad启动子甲基化状态的不显著变化一致。与此相反，其N-末端ATRX和PWWP结构域的删除废除了其阻遏功能。组蛋白去乙酰化酶2（Hdac 2）与T-Cad启动子的关联以及在用Hisperin抑制剂处理后Dnmt 3b介导的抑制的启动子活性的恢复表明组蛋白去乙酰化参与该过程。在PC 12细胞中，T-Cad异位表达后，NGF诱导的神经突生长以剂量依赖性方式受到抑制。免疫荧光研究表明，T-Cad的神经生长因子治疗后重新分布，从其浓度在轴突生长锥，而不是其定位在细胞-细胞接触区域的未分化细胞。这些结果证明了T-Cad在NGF介导的PC 12细胞向神经元表型分化中的新作用。
Previously we showed that DNA methyltransferase 3b (Dnmt3b) is required for nerve growth factor (NGF)- induced differentiation of PC12 cells to neuronal phenotype. The present study identified T-cadherin (T-Cad) as one of the targets of Dnmt3b by chromatin immunoprecipitation (ChIP) assay. Combined bisulfite restriction analysis and bisulfite sequencing showed that T- Cad promoter was sparsely methylated in PC12 cells. ChIP-CHOP analysis demonstrated that Dnmt3b is associated with T-Cad promoter irrespective of its methylation status. The mRNA and protein levels of T-Cad were markedly elevated in cells depleted of Dnmt3b by antisense or small interfering RNA. Suppression of T- Cad promoter activity by Dnmt3b was independent of its catalytic activity, which was consistent with the insignificant change in T-Cad promoter methylation status in Dnmt3b-depleted cells. In contrast, deletion of its N-terminal ATRX and PWWP domain abolished its repressor function. Association of histone deacetylase 2 (Hdac2) with T-Cad promoter and restoration of the promoter activity from Dnmt3b-mediated suppression upon treatment with Hdac inhibitor indicated involvement of histone deacetylation in this process. NGF-induced neurite outgrowth was inhibited in a dose dependent manner upon ectopic expression of T-Cad in PC12 cells. Immunofluorescence studies showed that T-Cad was redistributed upon NGF treatment, as evident from its concentration in axon growth cones as opposed to its localization at cell-cell contact region in undifferentiated cells. These results demonstrate a novel role of T-Cad in the NGF-mediated differentiation of PC12 cells to neuronal phenotype.