Moore William M, Brea Roberto J, Knittel Caroline H, Wrightsman Ellen, Hui Brandon, Lou Jinchao, Ancajas Christelle F, Best Michael D, Obara Christopher J, Devaraj Neal K, Budin Itay
Department of Chemistry and Biochemistry, University of California, San Diego, La Jolla, CA, USA.
Centro Interdisciplinar de Química e Bioloxía (CICA), Universidade da Coruña, A Coruña, Spain.
Nat Chem Biol. 2025 Sep 15. doi: 10.1038/s41589-025-02021-z.
The lipid composition of cells varies widely across organelles and between individual membrane leaflets. Transport proteins are thought to generate this heterogeneity, but measuring their functions in vivo has been hampered by limited tools for imaging lipids at relevant spatial resolutions. Here we present fluorogen-activating coincidence encounter sensing (FACES), a chemogenetic tool capable of quantitatively imaging subcellular lipid pools and reporting their transbilayer orientation in living cells. FACES combines bioorthogonal chemistry with genetically encoded fluorogen-activating proteins (FAPs) for reversible proximity sensing of conjugated molecules. We first apply this approach to identify roles for lipid transfer proteins that traffic phosphatidylcholine pools between the ER and mitochondria. We then show that transmembrane domain-containing FAPs can reveal the membrane asymmetry of multiple lipid classes in the trans-Golgi network and be used to investigate the mechanisms that generate it. Finally, we present that FACES can be applied to measure glycans and other molecule classes.
细胞的脂质组成在不同细胞器之间以及单个膜小叶之间差异很大。人们认为转运蛋白会产生这种异质性，但在体内测量它们的功能一直受到相关空间分辨率下脂质成像工具有限的阻碍。在这里，我们展示了荧光激活巧合相遇传感（FACES），这是一种化学遗传学工具，能够对亚细胞脂质池进行定量成像并报告它们在活细胞中的跨膜取向。FACES将生物正交化学与基因编码的荧光激活蛋白（FAPs）相结合，用于对共轭分子进行可逆的邻近传感。我们首先应用这种方法来确定在内质网和线粒体之间运输磷脂酰胆碱池的脂质转移蛋白的作用。然后我们表明，含跨膜结构域的FAPs可以揭示反式高尔基体网络中多种脂质类别的膜不对称性，并用于研究产生这种不对称性的机制。最后，我们展示了FACES可用于测量聚糖和其他分子类别。
