The characteristics of CTCF binding sequences contribute to enhancer blocking activity
The characteristics of CTCF binding sequences contribute to enhancer blocking activity
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DOI：
10.1101/2023.09.06.556325
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发表时间：
2023-09
期刊：
影响因子：
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通讯作者：
Felice H. Tsang;Rosa J. Stolper;Muhammad Hanifi;Lucy J. Cornell;Helena S. Francis;Benjamin Davies;D. Higgs;Mira T. Kassouf
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文献类型：
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作者：
Felice H. Tsang;Rosa J. Stolper;Muhammad Hanifi;Lucy J. Cornell;Helena S. Francis;Benjamin Davies;D. Higgs;Mira T. Kassouf
虽然编码增强子和启动子的元件已被相对较好地研究，但结合CCCTC结合因子（CTCF）的绝缘子元件的全谱特征相对较差。这部分是由于CTCF位点的基因组背景极大地影响了它们的作用和活性。在这里，我们已经开发了一个实验系统，以确定一致的大小，个别CTCF元件之间的增强子和启动子，从而减少分化过程中的基因表达的能力。重要的是，每个元件在相同的位置进行测试，从而最大限度地减少基因组背景的影响。我们发现CTCF元件阻断增强子-启动子活性的能力与这些元件的天然基因组位点上结合的CTCF或粘附素的量、进化保守的程度或它们与共有核心序列的相似性之间没有相关性。然而，我们已经表明，最强的增强子-启动子阻断剂包括一个先前描述的结合元件位于CTCF核心基序的上游。此外，我们还发现了其他未表征的DNaseI足迹，这些足迹位于可能影响功能的核心基序附近。我们已经开发了一种CTCF序列的测定方法，这将使研究人员能够以统一和公正的方式对CTCF元件进行细分。
While the elements encoding enhancers and promoters have been relatively well studied, the full spectrum of insulator elements which bind the CCCTC binding factor (CTCF), is relatively poorly characterised. This is partly due to the genomic context of CTCF sites greatly influencing their roles and activity. Here we have developed an experimental system to determine the ability of consistently sized, individual CTCF elements to interpose between enhancers and promoters and thereby reduce gene expression during differentiation. Importantly, each element is tested in the identical location thereby minimising the effect of genomic context. We found no correlation between the ability of CTCF elements to block enhancer-promoter activity with the amount of CTCF or cohesin bound at the natural genomic sites of these elements; the degree of evolutionary conservation; or their resemblance to the consensus core sequences. Nevertheless, we have shown that the strongest enhancer-promoter blockers include a previously described bound element lying upstream of the CTCF core motif. In addition, we found other uncharacterised DNaseI footprints located close to the core motif that may affect function. We have developed an assay of CTCF sequences which will enable researchers to sub-classify CTCF elements in a uniform and unbiased way.