A high-throughput small molecule screen identifies synergism between DNA methylation and Aurora kinase pathways for X reactivation
A high-throughput small molecule screen identifies synergism between DNA methylation and Aurora kinase pathways for X reactivation
复制标题
DOI：
10.1073/pnas.1617597113
复制
发表时间：
2016-12-13
影响因子：
11.1
通讯作者：
Lee, Jeannie T.
中科院分区：
文献类型：
--
作者：
Lessing, Derek;Dial, Thomas O.;Lee, Jeannie T.
X染色体失活是一种剂量补偿机制，其中雌性哺乳动物的两条X染色体之一在转录上沉默。一旦建立，失活X（Xi）的沉默是稳健的并且难以逆转突变。然而，Xi是一个拥有超过1,000个功能基因的水库，这些基因可能被用来治疗X连锁疾病。为了鉴定可以重新激活Xi的化合物，我们在小鼠成纤维细胞中使用Xi连接的GFP报告基因在自动化高含量筛选中筛选了类似于367，000个小分子。考虑到沉默的稳健性质，我们通过用DNA甲基转移酶抑制剂5-氮杂-2 '-脱氧胞苷（5azadC）“引发”细胞来敏化筛选。引发GFP活性的化合物包括VX 680、MLN 8237和5azadC，已知它们靶向Aurora激酶和DNA甲基化途径。我们证明了VX 680和5azadC以及MLN 8237和5azadC的组合协同上调Xi上的基因。因此，我们的工作确定了DNA甲基化和Aurora激酶途径之间的协同作用，这是可能的XI药理学重新激活的兴趣之一。
X-chromosome inactivation is a mechanism of dosage compensation in which one of the two X chromosomes in female mammals is transcriptionally silenced. Once established, silencing of the inactive X (Xi) is robust and difficult to reverse pharmacologically. However, the Xi is a reservoir of > 1,000 functional genes that could be potentially tapped to treat X-linked disease. To identify compounds that could reactivate the Xi, here we screened similar to 367,000 small molecules in an automated high-content screen using an Xi-linked GFP reporter in mouse fibroblasts. Given the robust nature of silencing, we sensitized the screen by "priming" cells with the DNA methyltransferase inhibitor, 5-aza-2'-deoxycytidine (5azadC). Compounds that elicited GFP activity include VX680, MLN8237, and 5azadC, which are known to target the Aurora kinase and DNA methylation pathways. We demonstrate that the combinations of VX680 and 5azadC, as well as MLN8237 and 5azadC, synergistically up-regulate genes on the Xi. Thus, our work identifies a synergism between the DNA methylation and Aurora kinase pathways as being one of interest for possible pharmacological reactivation of the Xi.