Hippo通路中转录激活因子YAP蛋白二聚体的分子机制研究

Hippo pathway was discovery as important signal pathway to regulate organ size. Transcription activator YAP is regulation effecter and output by Hippo pathway. In mammalian cell, when Hippo signal is activated, Protein kinase Mst2 and Lats1 phosphorylate YAP, and keep YAP in cytoplasm. Unphosphorylated YAP, When Hippo pathway was dysfunction, translocates into nucleus and interacts with transcription factor TEAD family. The complex YAP-TEAD promotes transcription of proliferative and pro-survival genes, such as CyclinE and Diap1, and induce cell proliferation, leads to hyperplasia and cancer formation. Clinic research found that overexpression of YAP protein or malfunction of Hippo pathway induce tumor and cancer, YAP was identified as an onco-protein and target for anti-cancer treatment and pre-diagnosis. Our preliminary results identified that YAP protein forms dimer in un-phosphorylated state from recombinant expression, and dimerized YAP interacts with Tead2 to form complex with 2:2 mole ratio. This project is going to focus on the molecular mechanism to induce YAP dimerization. We will search and identify which domain or molecule to mediate YAP dimerization by in-vitro and in-vivo experiment, and seek to further research of structure biology for YAP dimer. This proposal to focus on moleculare state and mechanism of YAP protein, will shed light on regulation mechanism of Hippo pathway, also provide new insight for anti-cancer treatment and diagnosis.

Hippo信号通路是近年来被发现的调控组织大小的重要信号通路。转录激活因子YAP是Hippo信号通路的效应分子和调控终端。当Hippo信号被激活后，蛋白激酶Mst2和Lats1磷酸化YAP，使其滞留于细胞质内。而非磷酸化的YAP蛋白转运至细胞核内，结合转录因子TEAD家族蛋白，介导目的基因的转录，引起细胞增殖而诱发组织肿大或肿瘤。临床医学研究发现，YAP蛋白过量表达诱发癌症，认为YAP蛋白是一种致癌蛋白，是癌症治疗和前期诊断的重要靶点。我们的前期研究发现重组的YAP蛋白以二聚体的形式存在并结合TEAD2蛋白。本项目将以YAP蛋白二聚体的形成原因及分子机制为主要研究目标，利用体外和体内实验，寻找YAP蛋白二聚体形成的分子成因和结构域，为进一步的结构生物学研究奠定基础。深入研究YAP蛋白的分子状态及分子机制，将有利于完整阐述Hippo信号通路的调控机制，为癌症治疗或前期诊断提供新角度。

Hippo信号通路是近年来被发现的调控组织大小的重要信号通路，在个体发育与肿瘤发生发展中发挥重要作用。转录激活因子YAP是Hippo信号通路的效应分子和调控终端，与转录因子Tead一起激活目的基因的转录与表达。医学研究发现，YAP蛋白过量表达诱发癌症，认为YAP蛋白是一种致癌蛋白，是癌症治疗和前期诊断的新靶点。深入研究YAP蛋白的分子状态及分子机制，将有利于完整阐述Hippo信号通路的调控机制，为癌症治疗或前期诊断提供新角度。我们的研究发现YAP蛋白以单体和二聚体两种构象方式存在，但二聚体的比例极低，约占10%的比例。进一步的研究发现TEAD与YAP二者形成复合物时的分子比例是1:1，并无二聚体形式存在。因此二聚体YAP可能无法结合转录因子TEAD蛋白，并可能拮抗YAP的转录激活活性。.肿瘤抑制因子NF2的缺失或突变，直接导致神经纤维瘤，基因敲除NF2导致小鼠在胚胎早期致死。近年来NF2被发现参与Hippo信号通路并处于上游位置，Merlin可激活Hippo信号通路的相关激酶进而抑制YAP蛋白的水平与活性，进而抑制肿瘤的发生与发展，但NF2抑制肿瘤的分子机制仍不清楚。我们的研究发现，NF2蛋白可以特异性的直接结合转录因子TEAD蛋白，并拮抗YAP-TEAD转录复合物的形成，从而抑制YAP作为转录增强因子的活性。进一步的研究发现NF2可介导Tead蛋白不依赖蛋白酶体的生物降解。该研究结果将从分子机制上揭示NF2作为肿瘤抑制因子的真正原因，为潜在的肿瘤治疗提供新靶点和新方向。