Abstract：
De novo variants in the Cytoplasmic FMR1-interacting protein 2 (CYFIP2) have been repeatedly associated with neurodevelopmental disorders and epilepsy, underscoring its critical role in brain development and function. While CYFIP2\'s role in regulating actin polymerization as part of the WAVE regulatory complex (WRC) is well-established, its additional molecular functions remain relatively unexplored. In this study, we performed unbiased quantitative proteomic analysis, revealing 278 differentially expressed proteins (DEPs) in the forebrain of Cyfip2 knock-out embryonic mice compared to wild-type mice. Unexpectedly, these DEPs, in conjunction with previously identified CYFIP2 brain interactors, included not only other WRC components but also numerous proteins associated with membraneless organelles (MLOs) involved in mRNA processing and translation within cells, including the nucleolus, stress granules, and processing bodies. Additionally, single-cell transcriptomic analysis of the Cyfip2 knock-out forebrain revealed gene expression changes linked to cellular stress responses and MLOs. We also observed morphological changes in MLOs in Cyfip2 knock-out brains and CYFIP2 knock-down cells under basal and stress conditions. Lastly, we demonstrated that CYFIP2 knock-down in cells, potentially through WRC-dependent actin regulation, suppressed the phosphorylation levels of the alpha subunit of eukaryotic translation initiation factor 2 (eIF2α), thereby enhancing protein synthesis. These results suggest a physical and functional connection between CYFIP2 and various MLO proteins and also extend CYFIP2\'s role within the WRC from actin regulation to influencing eIF2α phosphorylation and protein synthesis. With these dual functions, CYFIP2 may fine-tune the balance between MLO formation/dynamics and protein synthesis, a crucial aspect of proper mRNA processing and translation.
摘要：
细胞质FMR1相互作用蛋白2（CYFIP2）中的从头变异体与神经发育障碍和癫痫反复相关，强调其在大脑发育和功能中的关键作用。虽然CYFIP2作为WAVE调节复合物(WRC)的一部分在调节肌动蛋白聚合中的作用已广为人知，其额外的分子功能仍然相对未被探索。在这项研究中,我们进行了无偏的定量蛋白质组学分析，揭示了与野生型小鼠相比，Cyfip2敲除胚胎小鼠前脑中278种差异表达蛋白（DEP）。出乎意料的是,这些DEP,结合先前确定的CYFIP2大脑相互作用者，不仅包括其他WRC成分，而且还包括许多与细胞内mRNA加工和翻译相关的无膜细胞器（MLO）相关的蛋白质，包括核仁,应力颗粒，和处理机构。此外,Cyfip2敲除前脑的单细胞转录组分析显示，与细胞应激反应和MLO相关的基因表达变化。我们还观察到在基础和应激条件下Cyfip2敲除脑和CYFIP2敲除细胞中MLO的形态变化。最后,我们证明CYFIP2在细胞中敲低，可能通过WRC依赖性肌动蛋白调节，抑制真核翻译起始因子2（eIF2α）α亚基的磷酸化水平，从而增强蛋白质合成。这些结果表明CYFIP2与各种MLO蛋白之间存在物理和功能上的联系，并且还将CYFIP2在WRC中的作用从肌动蛋白调节扩展到影响eIF2α磷酸化和蛋白质合成。有了这些双重功能，CYFIP2可以微调MLO形成/动力学和蛋白质合成之间的平衡，正确的mRNA加工和翻译的关键方面。
