α微管蛋白翻译后修饰在胎盘滋养层细胞融合过程中的作用及与子痫前期发病的相关性

The placental multinucleated syncytiotrophoblast arises from the fusion of villous cytotrophoblasts, secreting important growth factors and hormones (human chorionic gonadotropin and progesterone, etc) and mediating gas and nutrient exchanges and waste transportation to maintain pregnancy. Dysregulation in syncytialization results in pregnancy-related disease, such as pre-eclampsia (PE). Cytoskeleton changes are critical to trophoblast fusion. Tubulin post-translational modifications regulate microtubule function and dynamics. However, their functions during syncytialization remain elusive. By utilizing a variety of primary cell or tissue models for trophoblast fusion, live-cell imaging and placenta-specific gene manipulation mouse models, this project sets out to investigate the function of α-tubulin post-translational modification such as detyrosination during syncytialization. By studying the abnormal expression of detyrosinated α-tubulin (detyr-α-tubulin) and tubulin tyrosine ligase (TTL) in pre-eclamptic placentae and the underlying molecular mechanisms, we aim to understand the function of post-translational modification of α-tubulin in pregnancy maintenance and the relevance of dysregulated detyr-α-tubulin to the pathology of PE. Information achieved from these studies will provide insights into elucidating the pathological mechanism of dysregulated placentation and developing therapeutic strategies of PE.

胎盘绒毛合体滋养层由细胞滋养层细胞融合形成，分泌多种生长因子与激素(人绒毛膜促性腺激素和孕酮等)，介导气体和营养物质交换及废物运输，对于正常妊娠的建立和维持至关重要；滋养层细胞合体化异常能引起子痫前期等妊娠疾病。细胞骨架的重构对于合体化过程的发生至关重要。微管蛋白的翻译后修饰是微管功能和动态变化的重要调控因素，其在滋养层细胞合体化过程中的作用尚不明确。本研究拟以多种合体化细胞模型、活细胞工作站和胎盘特异基因操作小鼠模型，研究α微管蛋白的去酪氨酸化等翻译后修饰对滋养层细胞合体化的影响和分子机制。通过比较正常孕妇和子痫前期患者胎盘中去酪氨酸化α微管蛋白和酪氨酸连接酶的差异并结合体外机理研究，确定α微管蛋白的翻译后修饰在正常妊娠维持中的作用机制，并阐明α微管蛋白的去酪氨酸化异常与子痫前期发病的相关性。以上研究将为阐释子痫前期的胎盘发育障碍机理和发展新的临床治疗方案提供理论依据。

人类胎盘滋养层细胞合胞体化是胎盘发育过程中最重要的事件之一。为了研究α微管蛋白翻译后修饰在胎盘滋养层细胞合胞体化中的作用机制，我们用多种手段标记细胞，建立了滋养层细胞融合过程的实时活细胞成像方法，研究了滋养层细胞融合过程及其微管动态变化。我们对BeWo细胞融合模型的转录组测序数据分析发现，融合后被微管蛋白酪氨酸连接酶（tubulin tyrosine ligase，TTL）负调控的去酪氨酸化α-微管蛋白（detyrosinated α-tubulin，detyr-α-tub）表达水平升高。在妊娠早期胎盘绒毛的细胞滋养层细胞中，TTL有相对较高的表达定位，但与融合标志分子不共定位。利用BeWo细胞融合模型的分子机制研究发现，滋养层细胞中TTL表达水平下调导致detyr-α-tub表达显著升高；而detyr-α-tub表达的恢复促进融合原蛋白Syncytin-2和间隙连接蛋白Connexin 43（Cx43）在细胞膜上富集，进而促进滋养层细胞融合。子痫前期患者胎盘细胞滋养层细胞自发融合能力受损，但可以通过下调TTL表达水平部分挽救。因此，本项目的研究结果表明，detyr-α-tub上调有助于融合蛋白在细胞膜上的聚集，从而促进滋养层细胞融合；detyr-α-tub不足引起合体化缺陷，并可能导致子痫前期。