Brumbaugh-Reed Ellen H, Aoki Kazuhiro, Toettcher Jared E
Department of Molecular Biology, Princeton University, Princeton NJ 08544.
Omenn-Darling Bioengineering Institute, Princeton University, Princeton NJ 08544.
bioRxiv. 2024 Jan 17:2024.01.16.575860. doi: 10.1101/2024.01.16.575860.
Biomolecular condensates are broadly implicated in both normal cellular regulation and disease. Consequently, several chemical biology and optogenetic approaches have been developed to induce phase separation of a protein of interest. However, few tools are available to perform the converse function-dissolving a condensate of interest on demand. Such a tool would aid in testing whether the condensate plays specific functional roles, a major question in cell biology and drug development. Here we report an optogenetic approach to selectively dissolve a condensate of interest in a reversible and spatially controlled manner. We show that light-gated recruitment of maltose-binding protein (MBP), a commonly used solubilizing domain in protein purification, results in rapid and controlled dissolution of condensates formed from proteins of interest. Our optogenetic MBP-based dissolution strategy (OptoMBP) is rapid, reversible, and can be spatially controlled with subcellular precision. We also provide a proof-of-principle application of OptoMBP, showing that disrupting condensation of the oncogenic fusion protein FUS-CHOP results in reversion of FUS-CHOP driven transcriptional changes. We envision that the OptoMBP system could be broadly useful for disrupting constitutive protein condensates to probe their biological functions.
生物分子凝聚物广泛参与正常细胞调节和疾病过程。因此，已经开发了几种化学生物学和光遗传学方法来诱导目标蛋白的相分离。然而，用于执行相反功能（即按需溶解目标凝聚物）的工具却很少。这样的工具将有助于测试凝聚物是否发挥特定的功能作用，这是细胞生物学和药物开发中的一个主要问题。在这里，我们报告了一种光遗传学方法，以可逆和空间可控的方式选择性地溶解目标凝聚物。我们表明，光门控募集麦芽糖结合蛋白（MBP），这是蛋白质纯化中常用的可溶结构域，可导致由目标蛋白形成的凝聚物快速且可控地溶解。我们基于光遗传学MBP的溶解策略（OptoMBP）快速、可逆，并且可以在亚细胞水平上进行空间控制。我们还提供了OptoMBP的原理验证应用，表明破坏致癌融合蛋白FUS-CHOP的凝聚会导致FUS-CHOP驱动的转录变化逆转。我们设想OptoMBP系统可广泛用于破坏组成型蛋白凝聚物以探究其生物学功能。
