Discovering human transcription factor physical interactions with genetic variants, novel DNA motifs, and repetitive elements using enhanced yeast one-hybrid assays
Discovering human transcription factor physical interactions with genetic variants, novel DNA motifs, and repetitive elements using enhanced yeast one-hybrid assays
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DOI：
10.1101/gr.248823.119
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发表时间：
2019-09
期刊：
影响因子：
7
通讯作者：
Shaleen Shrestha;J. Sewell;Clarissa S Santoso;Elena J. Forchielli;S. Carrasco Pro;Melissa Martinez;J. F. Fuxman Bass
中科院分区：
文献类型：
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作者：
Shaleen Shrestha;J. Sewell;Clarissa S Santoso;Elena J. Forchielli;S. Carrasco Pro;Melissa Martinez;J. F. Fuxman Bass
识别转录因子（TF）与非编码变体、未表征的DNA基序和重复基因组元件的结合在技术上和计算上都具有挑战性。目前的实验方法，如染色质免疫沉淀，一般测试一个TF的时间，和计算基序算法往往导致假阳性和阴性的预测。为了解决这些局限性，我们开发了一种基于增强型酵母单杂交测定的实验方法。该方法的第一种变体询问>1000个人类TF与重复DNA元件的结合，而第二种变体评估TF与单核苷酸变体、短插入和缺失（indel）以及新DNA基序的结合。使用这种方法，我们检测到75个TF，包括几个核激素受体和ETS因子，高度重复的Alu元素的结合。此外，我们确定了TF结合的癌症相关变化，包括涉及ETS TF的相互作用的获得和涉及KLF TF与TERT启动子中不同突变的相互作用的丧失，以及MYB与TAL1超增强子中18 bp indel的相互作用的获得。此外，我们还鉴定了与DNA酶足迹分析中鉴定的三个未表征的DNA基序结合的TF。我们预计，这些增强的酵母单杂交方法将扩大我们研究遗传变异和特征不足的基因组区域的能力。
Identifying transcription factor (TF) binding to noncoding variants, uncharacterized DNA motifs, and repetitive genomic elements has been technically and computationally challenging. Current experimental methods, such as chromatin immunoprecipitation, generally test one TF at a time, and computational motif algorithms often lead to false-positive and -negative predictions. To address these limitations, we developed an experimental approach based on enhanced yeast one-hybrid assays. The first variation of this approach interrogates the binding of >1000 human TFs to repetitive DNA elements, while the second evaluates TF binding to single nucleotide variants, short insertions and deletions (indels), and novel DNA motifs. Using this approach, we detected the binding of 75 TFs, including several nuclear hormone receptors and ETS factors, to the highly repetitive Alu elements. Further, we identified cancer-associated changes in TF binding, including gain of interactions involving ETS TFs and loss of interactions involving KLF TFs to different mutations in the TERT promoter, and gain of a MYB interaction with an 18-bp indel in the TAL1 superenhancer. Additionally, we identified TFs that bind to three uncharacterized DNA motifs identified in DNase footprinting assays. We anticipate that these enhanced yeast one-hybrid approaches will expand our capabilities to study genetic variation and undercharacterized genomic regions.