Communication Routes in ARID Domains between Distal Residues in Helix 5 and the DNA-Binding Loops

ARID is a DNA-binding domain involved in several transcriptional regulatory processes, including cell-cycle regulation and embryonic development. ARID domains are also targets of the Human Cancer Protein Interaction Network. Little is known about the molecular mechanisms related to conformational changes in the family of ARID domains. Thus, we have examined their structural dynamics to enrich the knowledge on this important family of regulatory proteins. In particular, we used an approach that integrates atomistic simulations and methods inspired by graph theory. To relate these properties to protein function we studied both the free and DNA-bound forms. The interaction with DNA not only stabilizes the conformations of the DNA-binding loops, but also strengthens pre-existing paths in the native ARID ensemble for long-range communication to those loops. Residues in helix 5 are identified as critical mediators for intramolecular communication to the DNA-binding regions. In particular, we identified a distal tyrosine that plays a key role in long-range communication to the DNA-binding loops and that is experimentally known to impair DNA-binding. Mutations at this tyrosine and in other residues of helix 5 are also demonstrated, by our approach, to affect the paths of communication to the DNA-binding loops and alter their native dynamics. Overall, our results are in agreement with a scenario in which ARID domains exist as an ensemble of substates, which are shifted by external perturbation, such as the interaction with DNA. Conformational changes at the DNA-binding loops are transmitted long-range by intramolecular paths, which have their heart in helix 5.

ARID是一类参与多种转录调控过程的DNA结合结构域（DNA-binding domain），其调控通路涵盖细胞周期调控与胚胎发育。ARID结构域同时也是人类癌症蛋白质相互作用网络（Human Cancer Protein Interaction Network）的作用靶点。目前学界对ARID家族蛋白质构象变化相关的分子机制仍所知甚少。为此，我们针对其结构动力学展开了系统研究，以期丰富这一重要调控蛋白家族的相关认知。具体而言，我们采用了整合全原子模拟（atomistic simulations）与图论启发方法的研究策略。为将这些结构特性与蛋白质功能相关联，我们同时对游离态与DNA结合态的ARID结构域进行了分析。与DNA的相互作用不仅能够稳定DNA结合环的构象，还能强化天然ARID构象集合中通往这些结合环的长距离通信通路。研究发现，螺旋5（helix 5）中的残基是介导DNA结合区域分子内通信的关键因子。特别地，我们鉴定出一个远端酪氨酸残基，其在通往DNA结合环的长距离通信中发挥关键作用，且已有实验证实该位点突变会损害DNA结合能力。通过本研究的分析方法，我们还证实，该酪氨酸残基与螺旋5中其他残基的突变，会破坏通往DNA结合环的通信通路并改变其天然动力学特性。总体而言，我们的研究结果支持如下理论模型：ARID结构域以多种构象亚态的集合形式存在，这类亚态会因外部扰动（如与DNA的相互作用）发生偏移；DNA结合环的构象变化通过以螺旋5为核心的分子内通路实现长距离传递。