精氨酸甲基化（ArgMet）作为一种翻译后修饰，在RNA加工、转录调控、信号转导、DNA修复、细胞凋亡和液-液相分离（LLPS）中发挥着至关重要的作用。由于精氨酸甲基化与癌症发病机制和进展相关，因此蛋白质精氨酸甲基转移酶作为抗癌治疗的靶标引起了人们的兴趣。尽管已经进行了大量的工作来阐明精氨酸甲基化调节的（病理）生理机制，但仍然缺乏在活细胞中以高时空分辨率可视化精氨酸甲基化的工具。为了解决这一未满足的需求，我们生成了一种对 ArgMet 敏感的基因编码、基于福斯特共振能量转移 (FRET) 的生物传感器，称为 GEMS，能够定量实时监测 ArgMet 动态。我们通过使用不同的 ArgMet 结合域、富含精氨酸-甘氨酸的区域并调整生物传感器内的接头来优化这些生物传感器以提高其性能。使用一组哺乳动物细胞系和调节剂，我们证明了 GEMS 在以单细胞和时间分辨率监测精氨酸甲基化变化方面的适用性。 GEMS 可以促进体外筛选，以发现潜在的蛋白质精氨酸甲基转移酶抑制剂，并将有助于更好地了解与分化、发育和疾病相关的 ArgMet 调节。版权所有 © 2024 作者。由 Elsevier B.V. 出版。保留所有权利。

Arginine methylation (ArgMet), as a post-translational modification, plays crucial roles in RNA processing, transcriptional regulation, signal transduction, DNA repair, apoptosis and liquid-liquid phase separation (LLPS). Since arginine methylation is associated with cancer pathogenesis and progression, protein arginine methyltransferases have gained interest as targets for anti-cancer therapy. Despite considerable process made to elucidate (patho)physiological mechanisms regulated by arginine methylation, there remains a lack of tools to visualize arginine methylation with high spatiotemporal resolution in live cells. To address this unmet need, we generated an ArgMet-sensitive genetically encoded, Förster resonance energy transfer-(FRET) based biosensor, called GEMS, capable of quantitative real-time monitoring of ArgMet dynamics. We optimized these biosensors by using different ArgMet-binding domains, arginine-glycine-rich regions and adjusting the linkers within the biosensors to improve their performance. Using a set of mammalian cell lines and modulators, we demonstrated the applicability of GEMS for monitoring changes in arginine methylation with single-cell and temporal resolution. The GEMS can facilitate the in vitro screening to find potential protein arginine methyltransferase inhibitors and will contribute to a better understanding of the regulation of ArgMet related to differentiation, development and disease.Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.