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A High-Throughput Genome-Integrated Assay Reveals Spatial Dependencies Governing Tcf7l2 Binding.
Cell Systems ( IF 9.3 ) Pub Date : 2020-09-09 , DOI: 10.1016/j.cels.2020.08.004
Tomasz Szczesnik 1 , Lendy Chu 2 , Joshua W K Ho 3 , Richard I Sherwood 4
Affiliation  

Predicting where transcription factors bind in the genome from their in vitro DNA-binding affinity is confounded by the large number of possible interactions with nearby transcription factors. To characterize the in vivo binding logic for the Wnt effector Tcf7l2, we developed a high-throughput screening platform in which thousands of synthesized DNA phrases are inserted into a specific genomic locus, followed by measurement of Tcf7l2 binding by DamID. Using this platform at two genomic loci in mouse embryonic stem cells, we show that while the binding of Tcf7l2 closely follows the in vitro motif-binding strength and is influenced by local chromatin accessibility, it is also strongly affected by the surrounding 99 bp of sequence. Through controlled sequence perturbation, we show that Oct4 and Klf4 motifs promote Tcf7l2 binding, particularly in the adjacent 50 bp and oscillating with a 10.8-bp phasing relative to these cofactor motifs, which matches the turn of a DNA helix.



中文翻译:

高通量基因组整合分析揭示了控制 Tcf7l2 结合的空间依赖性。

从它们的体外DNA 结合亲和力预测转录因子在基因组的结合位置被大量可能与附近转录因子的相互作用所混淆。为了表征Wnt 效应子 Tcf7l2 的体内结合逻辑,我们开发了一个高通量筛选平台,其中将数千个合成的 DNA 短语插入特定基因组位点,然后通过 DamID 测量 Tcf7l2 结合。在小鼠胚胎干细胞中的两个基因组位点使用这个平台,我们表明虽然 Tcf7l2 的结合密切遵循体外基序结合强度受局部染色质可及性的影响,也受周围 99 bp 序列的强烈影响。通过受控序列扰动,我们表明 Oct4 和 Klf4 基序促进了 Tcf7l2 结合,尤其是在相邻的50 bp 并以相对于这些辅因子基序的 10.8 bp 相位振荡,这与 DNA 螺旋的转向相匹配。

更新日期:2020-09-23
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