Sugino Y, Misawa A, Inoue J, Kitagawa M, Hosoi H, Sugimoto T, Imoto I, Inazawa J
Department of Molecular Cytogenetics, Medical Research Institute and School of Biomedical Science, Tokyo Medical and Dental University, Tokyo, Japan.
Oncogene. 2007 Nov 22;26(53):7401-13. doi: 10.1038/sj.onc.1210550. Epub 2007 May 28.
We previously identified a cluster of prostanoid receptor genes, prostaglandin D2 receptor (PTGDR) and prostaglandin E receptor 2 (PTGER2), as possible targets for DNA methylation in advanced types of neuroblastoma (NB) using bacterial artificial chromosome array-based methylated CpG island amplification method. Among them, in this study, we found that PTGER2 was frequently silenced in NB cell lines, especially in those with MYCN amplification, through epigenetic mechanisms. In NB cell lines, DNA methylation pattern within a part of CpG island was inversely correlated with PTGER2 expression, and histone H3 and H4 deacetylation and histone H3 lysine 9 methylation within the putative promoter region were more directly correlated with silencing of this gene. Methylation of PTGER2 was observed more frequently in advanced-type of primary NBs compared with early-stage tumors. Growth of NB cells lacking endogenous PTGER2 expression was inhibited by restoration of the gene product by transient and stable transfection. A PTGER2-selective agonist, butaprost, increased intracellular cyclic adenosine monophosphate (cAMP) level, inhibited cell growth and induced apoptosis of NB cells stably expressing exogenous PTGER2. 8-Bromo-cAMP also inhibited growth of NB cells lacking PTGER2 expression, but not cells expressing this gene. Taken together, it is suggested that NB cells may lose responsiveness to PTGER2-mediated growth inhibition/apoptosis through epigenetic silencing of PTGER2 and/or disruption of downstream cAMP-dependent pathway during the neuroblastomagenesis.
我们之前使用基于细菌人工染色体阵列的甲基化CpG岛扩增方法，确定了一组前列腺素受体基因，即前列腺素D2受体（PTGDR）和前列腺素E受体2（PTGER2），它们可能是晚期神经母细胞瘤（NB）中DNA甲基化的靶点。其中，在本研究中，我们发现PTGER2在NB细胞系中经常通过表观遗传机制沉默，尤其是在那些MYCN扩增的细胞系中。在NB细胞系中，部分CpG岛内的DNA甲基化模式与PTGER2表达呈负相关，而假定启动子区域内的组蛋白H3和H4去乙酰化以及组蛋白H3赖氨酸9甲基化与该基因的沉默更直接相关。与早期肿瘤相比，在晚期原发性NB中更频繁地观察到PTGER2的甲基化。通过瞬时和稳定转染恢复基因产物可抑制缺乏内源性PTGER2表达的NB细胞的生长。一种PTGER2选择性激动剂布他前列素可提高细胞内环磷酸腺苷（cAMP）水平，抑制细胞生长并诱导稳定表达外源性PTGER2的NB细胞凋亡。8-溴-cAMP也抑制缺乏PTGER2表达的NB细胞的生长，但不抑制表达该基因的细胞。综上所述，提示NB细胞在神经母细胞瘤发生过程中可能通过PTGER2的表观遗传沉默和/或下游cAMP依赖性途径的破坏，失去对PTGER2介导的生长抑制/凋亡的反应性。
