Synthetic enhancer design by in silico compensatory evolution reveals flexibility and constraint in cis-regulation.
Synthetic enhancer design by in silico compensatory evolution reveals flexibility and constraint in cis-regulation.
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DOI：
10.1186/s12918-017-0485-2
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发表时间：
2017-11-29
影响因子：
--
通讯作者：
Reinitz J
中科院分区：
文献类型：
--
作者：
Barr KA;Martinez C;Moran JR;Kim AR;Ramos AF;Reinitz J
结合特定化学机制的模型已经成功地描述了果蝇发育增强子作为潜在转录因子结合基序的功能的活性。尽管如此，从其组成部分重建增强子所需的最小机制集尚不清楚。合成生物学提供了测试已知机制是否足以描述增强子活性的潜力，以及揭示对基序数量、顺序和间距的限制。使用的功能模型和在硅片补偿进化，我们产生了推定的合成甚至跳过条纹2增强子与不同程度的相似性的天然增强子。这些元件代表了天然stripe 2增强子向从头设计的两种合成增强子的进化轨迹。在第一个轨迹中，即使在超过三分之一的结合位点丢失后，空间调控的表达仍得以维持。在第二种情况下，与天然元素高度相似的序列并不驱动表达，但高度发散的序列约为最小条纹2增强子长度的一半，驱动了10倍的表达。此外，Zelda或Stat 92 E基序的同型簇，而不是Bicoid，驱动发育中的胚胎中的表达。在这里，我们提出了一个基因调控的功能模型来测试已知的转录因子及其相互作用在多大程度上解释了果蝇甚至跳过条纹2增强子的活性。第一个轨迹的初步成功表明，基因调控模型解释了条纹2增强子的大部分功能。表达偏离预测的情况表明未描述的因素可能起调节表达的作用。我们还发现，激活驱动的Bicoid和Hunchback对结合基序的空间组织高度敏感。相比之下，Zelda和Stat 92 E驱动简单同型簇的表达，表明这些因子驱动的激活较少受到限制。总的来说，这项工作中产生的40个序列为构建未来的基因调控模型提供了强大的训练集。本文的在线版本（doi：10.1186/s12918-017-0485-2）包含补充材料，可供授权用户使用。
Models that incorporate specific chemical mechanisms have been successful in describing the activity of Drosophila developmental enhancers as a function of underlying transcription factor binding motifs. Despite this, the minimum set of mechanisms required to reconstruct an enhancer from its constituent parts is not known. Synthetic biology offers the potential to test the sufficiency of known mechanisms to describe the activity of enhancers, as well as to uncover constraints on the number, order, and spacing of motifs. Using a functional model and in silico compensatory evolution, we generated putative synthetic even-skipped stripe 2 enhancers with varying degrees of similarity to the natural enhancer. These elements represent the evolutionary trajectories of the natural stripe 2 enhancer towards two synthetic enhancers designed ab initio. In the first trajectory, spatially regulated expression was maintained, even after more than a third of binding sites were lost. In the second, sequences with high similarity to the natural element did not drive expression, but a highly diverged sequence about half the length of the minimal stripe 2 enhancer drove ten times greater expression. Additionally, homotypic clusters of Zelda or Stat92E motifs, but not Bicoid, drove expression in developing embryos. Here, we present a functional model of gene regulation to test the degree to which the known transcription factors and their interactions explain the activity of the Drosophila even-skipped stripe 2 enhancer. Initial success in the first trajectory showed that the gene regulation model explains much of the function of the stripe 2 enhancer. Cases where expression deviated from prediction indicates that undescribed factors likely act to modulate expression. We also showed that activation driven Bicoid and Hunchback is highly sensitive to spatial organization of binding motifs. In contrast, Zelda and Stat92E drive expression from simple homotypic clusters, suggesting that activation driven by these factors is less constrained. Collectively, the 40 sequences generated in this work provides a powerful training set for building future models of gene regulation. The online version of this article (doi:10.1186/s12918-017-0485-2) contains supplementary material, which is available to authorized users.
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影响因子：
2.4
作者：
Janssens, H;Kosman, D;Reinitz, J
通讯作者：
Reinitz, J
影响因子：
4.5
作者：
Hare, Emily E.;Peterson, Brant K.;Iyer, Venky N.;Meier, Rudolf;Eisen, Michael B.
通讯作者：
Eisen, Michael B.
影响因子：
9.9
作者：
Fakhouri, Walid D.;Ay, Ahmet;Sayal, Rupinder;Dresch, Jacqueline;Dayringer, Evan;Arnosti, David N.
通讯作者：
Arnosti, David N.
影响因子：
30.8
作者：
Janssens, Hilde;Hou, Shuling;Reinitz, John
通讯作者：
Reinitz, John
影响因子：
4.5
作者：
Kim, Ah-Ram;Martinez, Carlos;Reinitz, John
通讯作者：
Reinitz, John