应激颗粒核心组分G3BP1,TTP对应激颗粒组装去组装的调控机制研究

Stress granule, a membrane-less organelle. Under stress, the initiation of translation arrested。Suppressed mRNA moves out from nucleus and into the cytoplasm to form stress granule. Stress granules are ribonucleoprotein granule formed by the RNA and proteins. This is liquid-liquid phase separation. When the stress signal is released, the stress granule will dis-assemble, and their components will be degraded. If for some reason the components of the stress granules accumulated, the phase transition will occur, and amyloid¬ like filaments will occur. If this happens in the nerve system, may cause neurodegenerative diseases. The core components of stress granule are often associated with neurodegenerative diseases. Our research will focus on the molecular mechanism of the regulation of stress granule assembly and dis-assembly by RNA-binding protein G3BP-1, TTP and related proteins. There is growing evidence that the multivalent macromolecular interactions is an important organizing principle for phase separation. In this study, the multivalent interaction of the core components of stress particles in the formation of stress particles will be accurately determined by multi-dimensional nuclear magnetic resonance spectroscopy. Post-translational modifications (PTMs) in low complexity (LC) and intrinsically disordered (ID) regions of SG-nucleating proteins have been proposed to regulate SG assembly and the molecular mechanism for these regulation will be studied. The process and mechanism of liquid-solid phase transition of major component of stress granules will be studied by multi-dimensional solid-state NMR experiments and electrochemical scanning tunneling microscopy.

应急颗粒，是一种无膜细胞器，在应激条件下，翻译受阻的mRNA与它结合的蛋白质就会出核，到细胞质中组装成应激颗粒。应激颗粒是RNA与蛋白质结合形成的核糖核蛋白质颗粒，呈液滴状。这就是液-液相分离。应激信号解除后应激颗粒会解聚，其组分会被降解。若因某种原因应激颗粒组分大量积累就会发生相变，纤维化，这若发生在神经细胞中，就可能造成神经退行性疾病。本项申请将集中研究应激颗粒核心组分，RNA结合蛋白质G3BP-1,TTP及相关蛋白质对应激颗粒组装及解聚调控的分子机制。越来越多的证据显示蛋白质之间、蛋白质与RNA间或RNA之间的多价相互作用是导致的相分离，形成无膜细胞器的一个重要机理。本项研究将用多维核磁共振技术精准测定应激颗粒核心组分在形成应激颗粒核心过程的多价相互作用，研究蛋白质翻译后修饰对应激颗粒形成或解聚的调控，用多维固体核磁实验，和电化学扫描隧道显微镜研究应激颗粒液-固相变的过程和机理。

本项申请集中研究应激颗粒核心组分，RNA结合蛋白质G3BP-1,TTP及相关蛋白质对应激颗粒组装及解聚调控的分子机制。.1..研究应激颗粒中的核心组分G3BP1不对称二甲基化精氨酸修饰与它的识别蛋白TDRD3的相互作用，阐述翻译后修饰（精氨酸甲基化）对应激颗粒的组装与解聚调控的分子机理。.TDRD3是不对称二甲基化精氨酸的识别蛋白，它定位于应激颗粒，它在应激颗粒组装中的功能尚不清楚。我们通过研究发现TDRD3与不对称二甲基化精氨酸修饰的G3BP1结合，介导甲基化G3BP1与RNA的相互作用，形成G3BP1-TDRD3-RNA的互作网络，甲基化的G3BP1可招募多个TDRD3，这增加了互作价态，促进了生物大分子凝聚体的液液相分离与应激颗粒的组装。在此基础上我们还开发了一个二价态抑制剂分子，它破坏了TDRD3与甲基化G3BP1的互作，抑制了G3BP1-TDRD3-RNA体系的液液相分离，进而减少了砷酸盐诱导的应激颗粒的形成，从而增加了肿瘤细胞对化疗药物的敏感性。这工作揭示了TDRD3在应激颗粒组装中的作用，展示了通过靶向生物大分子凝聚体治疗癌症的潜力。最近投送Nature communication, (reference number: NCOMMS-23-04308-T)，正在审稿中。.2..用液体核磁共振方法，研究应激颗粒核心组分TTP锌指结构域RNA的识别。目前我们已经完成TTP锌指结构域及的核磁谱峰的指认，并探究了TTP的N端和C端无序结构域，以及RNA对TTP相分离能力的影响。