Evaluating the effect of splicing mutations on isoform networks in autism

 描述(申请人提供)：大量导致精神障碍的突变已被确认。然而，这些突变导致这些疾病的机制仍不清楚。整合大型和异质数据集的计算方法将在预测精神疾病突变背后的分子机制方面发挥关键作用。在我们最近的研究中(Lin等人，Neuron，2015)，我们展示了时空蛋白质相互作用网络如何帮助识别16p11.2拷贝数变异缺失和复制携带者中可能参与调节大脑大小的路径。在这里，我们建议使用类似的方法来构建异构体水平的共表达和蛋白质相互作用网络，以预测自闭症谱系障碍(ASD)患者的从头剪接位点突变对功能的影响。目前在ASD患者中发现了80多个剪接位点新生突变，但这些突变中的任何一个都没有建立单一的致病机制。我们假设，异形体水平的网络将为我们提供一幅更详细、更现实的图像，了解大脑中ASD突变所破坏的过程。为了验证这一假设，我们提出了一种综合的方法，结合网络生物学、CRISPR/CAS技术、转录组和蛋白质组方法来预测和验证剪接位点突变对人类(IPSCs)和ASD动物模型中细胞和分子通路的影响。该项目的最终目标是预测和验证受从头ASD剪接位点突变影响的特定通路。我们将通过以下具体目标来实现这一目标：(1)建立和分析大脑共表达和物理相互作用的同形水平蛋白质网络；(2)将ASD突变映射到同形水平网络以预测其功能影响；(3)在神经元和动物模型中使用CRISPR/Cas技术验证中断的网络和通路。这项拟议的研究将发现并表征被新剪接位点ASD突变破坏的细胞和分子过程。这项研究中确定的通路可能是未来治疗干预的重要新靶点。

 DESCRIPTION (provided by applicant): A large number of mutations contributing to psychiatric disorders have been identified. However, the mechanisms by which these mutations cause these diseases are still unknown. Computational approaches that integrate large and heterogeneous datasets will play crucial role in predicting molecular mechanisms behind psychiatric disease mutations. In our recent study (Lin et al, Neuron, 2015) we demonstrated how spatio-temporal protein interaction networks could help to identify a pathway that is likely involved in regulating brain size in the 16p11.2 copy number variant deletion and duplication carriers. Here, we propose to use similar approaches to construct the isoform-level co-expression and protein interaction networks for predicting functional impact of the de novo splice site mutations from the patients with autism spectrum disorder (ASD). More than 80 splice site de novo mutations are currently identified in the ASD patients, but not a single disease mechanism is established for any of these mutations. We hypothesize that isoform-level networks will provide us with a more detailed and realistic picture of the processes that are disrupted by the ASD mutations in the brain. To test this hypothesis, we propose an integrative approach that combines network biology, CRISPR/Cas technology, transcriptomic and proteomic methods to predict and validate the impact of splice site mutations on cellular and molecular pathways in the human (iPSCs) and animal models of ASD. The ultimate goal of this project is to predict and validate specific pathways that are impacted by the de novo ASD splice site mutations. We will achieve this goal through the following specific aims: (1) Build and analyze isoform-level networks of brain co-expressed and physically interacting proteins; (2) Map de novo ASD mutations onto isoform-level networks to predict their functional impact; (3) Validate the disrupted networks and pathways using CRISPR/Cas technology in neuronal and animal models. The proposed study will discover and characterize cellular and molecular processes that are disrupted by the de novo splice site ASD mutations. The pathways identified in this study could represent important new targets for future therapeutic intervention.