Acta Neuropathologica: C11orf95-RELA对幕上室管膜瘤的三维表观基因组进行了重新编程

2020-09-16 MedSci原创 MedSci原创

幕上室管膜瘤(ST-EPN)是一种主要见于儿童的恶性脑肿瘤。根据肿瘤部位,它可分为幕上型、幕下型和脊髓型三类。目前尚无有效的化疗方案,其治疗仅限于手术加或不加辅助放疗。

幕上室管膜瘤(ST-EPN)是一种主要见于儿童的恶性脑肿瘤。根据肿瘤部位,它可分为幕上型、幕下型和脊髓型三类。目前尚无有效的化疗方案,其治疗仅限于手术加或不加辅助放疗。这是因为室管膜瘤的很大一部分是在幼儿身上发现的,放射治疗并不可取,因为它会对发育中的大脑产生有害影响。因此,目前迫切需要发展基于基础生物学的靶向治疗。自2014年以来,已知染色体内融合C11orf95-RELA是ST-EPN的致癌驱动因素,在大约70%的ST-EPN中,C11orf95 RELA融合是最常见的结构变异。C11orf95-RELA转化的神经干细胞能够在小鼠体内形成脑肿瘤。然而,其在肿瘤发生过程中的作用在分子水平上还不清楚。

在本文中,作者团队发现融合蛋白的C11orf95成分决定DNA结合活性,而RELA成分则是驱动室管膜瘤相关基因表达所必需的。利用ChIP-seq和HiChIP方法进行的表观遗传学研究表明,C11orf95 RELA调节染色质状态并介导染色质相互作用,导致室管膜瘤细胞转录重编程。本文的发现为C11orf95 RELA融合的分子机制提供了重要的表征,并为C11orf95 RELA亚型室管膜瘤的潜在治疗靶点提供了依据。研究结果表明,与C11orf95 RELA室管膜瘤是由融合的RELA成分驱动的假设相反,融合蛋白与DNA的结合亲和力是由C11orf95成分决定的。RELA组分的作用是稳定融合蛋白的DNA结合,并提供其激活域来驱动靶基因的表达。

方法:HEK293T及其衍生的G16-2、G16-3、G16-4细胞以及BXD-1425-EPN细胞在杜尔贝科改良的Eagle培养基(Sigma)中培养,该培养基中含有10%胎牛血清(Lonza)、4%谷氨酰胺(Gibco)、1%丙酮酸钠(Gibco)和青霉素链霉素(Gibco)。培养箱条件为37°C和5%CO2。1μg/ml多西环素处理2天,诱导G16-2、G16-3和G16-4细胞中蛋白的过度表达。为了构建G16-2、G16-3和G16-4细胞系,在转染前一天,将HEK293T细胞以每孔1.5×105个细胞的密度接种在12孔板中。在转化之前,将培养基改为含有10μg/ml聚凝胺的培养基,每孔1 ml。将每种病毒各250μl添加到每个孔中并培养过夜。转染后24小时更换培养基。转染4天后,细胞分裂。转染后5天,将含有抗生素(2μg/ml嘌呤霉素)的培养基添加到一个复制板的每个孔中。在下游实验中使用抗生素前,抗生素选择至少持续了2周。

总之,虽然本研究首次提出了C11orf95-RELA融合蛋白诱导的遗传程序的表观遗传学和转录学特征,未来对其他ST-EPN-RELA细胞系和原发性肿瘤的研究将进一步确立核心程序以及亚型差异,这对于开发这种毁灭性癌症的精确和个性化治疗至关重要。

Zhu, J.J., Jillette, N., Li, X. et al. C11orf95-RELA reprograms 3D epigenome in supratentorial ependymoma. Acta Neuropathol (2020). 

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    2021-03-02 yb6560
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    2020-09-28 windight
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