A transcription factor network represses CFTR gene expression in airway epithelial cells
A transcription factor network represses CFTR gene expression in airway epithelial cells
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DOI：
10.1042/bcj20180044
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发表时间：
2018-04-16
影响因子：
4.1
通讯作者：
Harris, Ann
中科院分区：
文献类型：
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作者：
Mutolo, Michael J.;Leir, Shih-Hsing;Harris, Ann
囊性纤维化跨膜传导调节因子（CFTR）基因突变导致遗传性疾病囊性纤维化（CF）。肺部疾病是CF发病的主要原因，尽管CFTR表达水平在气道上皮中显著低于胰管和肠上皮，其也显示CF的功能受损。最近开发的用于CF的小分子疗法对于一种特定的CFTR突变非常成功，并且对其他突变具有积极影响。然而，气道中CFTR转录物的低丰度限制了药物纠正缺陷底物的机会。CFTR基因座的转录机制的阐明主要集中在基因内和基因间的组织特异性增强子及其激活的反式因子。在这里，我们调查是否在气道上皮细胞中的低CFTR水平的结果，直接向该位点的抑制蛋白的招聘。使用siRNA筛选来耗尽Calu-3肺上皮细胞中类似于1500种转录因子（TF）和相关调节蛋白，我们鉴定了近40种因子，其在耗尽后使CFTR mRNA水平升高超过2倍。这些TF的一个子集在原代人支气管上皮细胞中得到验证。在CFTR气道表达的最强阻遏物中，Kruppel样因子5和Ets同源因子，两者在气道上皮中具有关键作用。这些因子都被募集到距CFTR启动子约35 kb处的气道选择性顺式调节元件，它们的耗尽改善了CFTR的产生和功能，从而定义了用于增强CFTR的新的治疗靶标。
Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene cause the inherited disorder cystic fibrosis (CF). Lung disease is the major cause of CF morbidity, though CFTR expression levels are substantially lower in the airway epithelium than in pancreatic duct and intestinal epithelia, which also show compromised function in CF. Recently developed small molecule therapeutics for CF are highly successful for one specific CFTR mutation and have a positive impact on others. However, the low abundance of CFTR transcripts in the airway limits the opportunity for drugs to correct the defective substrate. Elucidation of the transcriptional mechanisms for the CFTR locus has largely focused on intragenic and intergenic tissue-specific enhancers and their activating trans-factors. Here, we investigate whether the low CFTR levels in the airway epithelium result from the recruitment of repressive proteins directly to the locus. Using an siRNA screen to deplete similar to 1500 transcription factors (TFs) and associated regulatory proteins in Calu-3 lung epithelial cells, we identified nearly 40 factors that upon depletion elevated CFTR mRNA levels more than 2-fold. A subset of these TFs was validated in primary human bronchial epithelial cells. Among the strongest repressors of airway expression of CFTR were Kruppel-like factor 5 and Ets homologous factor, both of which have pivotal roles in the airway epithelium. Depletion of these factors, which are both recruited to an airway-selective cis-regulatory element at -35 kb from the CFTR promoter, improved CFTR production and function, thus defining novel therapeutic targets for enhancement of CFTR.