Johannes Gutenberg-Universität (JGU), Biocenter, Institute of Molecular Physiology, Mainz, Germany.
Graduate School of Systemic Neurosciences (GSN), Planegg-Martinsried, Germany.
Methods Mol Biol. 2023;2563:325-339. doi: 10.1007/978-1-0716-2663-4_16.
In the last years, RNA-binding proteins (RBPs) have been highlighted for their capacity to undergo liquid-liquid phase separation (LLPS). Aberrant phase transitions of RBPs from a liquid to a solid state are believed to underlie the formation of pathological RBP aggregates in several neurodegenerative diseases. Both in the physiological and the disease state, RBPs are often decorated with diverse posttranslational modifications (PTMs) that can influence the phase separation behavior, the physiological function, and the pathological behavior of the RBP. Here we describe two simple methods, sedimentation assays in vitro and in cells, that allow the analysis of RBP solubility as a measure of RBP phase separation in the absence or presence of a certain PTM.
在过去的几年中，RNA 结合蛋白 (RBP) 因其能够进行液-液相分离 (LLPS) 的能力而受到关注。人们认为，RBP 从液相到固相的异常相转变是几种神经退行性疾病中病理性 RBP 聚集形成的基础。在生理和疾病状态下，RBP 通常被多种翻译后修饰 (PTM) 所修饰，这些修饰可以影响 RBP 的相分离行为、生理功能和病理行为。在这里，我们描述了两种简单的方法，即体外和细胞中的沉淀分析，可用于分析 RBP 的可溶性，作为在不存在或存在特定 PTM 的情况下 RBP 相分离的衡量标准。
