Complementary probes reveal that phosphatidylserine is required for the proper transbilayer distribution of cholesterol
Complementary probes reveal that phosphatidylserine is required for the proper transbilayer distribution of cholesterol
复制标题
DOI：
10.1242/jcs.164715
复制
发表时间：
2015-04-01
影响因子：
4
通讯作者：
Fairn, Gregory D.
中科院分区：
文献类型：
--
作者：
Maekawa, Masashi;Fairn, Gregory D.
胆固醇是后生动物细胞膜的重要组成部分，它有助于维持质膜的结构完整性和流动性。在这里，我们开发了一种胆固醇生物传感器，称为D4 H，基于产气荚膜梭菌θ-毒素的第四个结构域，其识别质膜和细胞器的胞质小叶中的胆固醇。D4 H探针在胆固醇提取后和细胞胆固醇运输扰动后与质膜分离。当与重组版本的生物传感器组合使用时，我们表明质膜磷脂酰丝氨酸是必不可少的保留在质膜的胞质小叶中的胆固醇。体外实验表明，1-硬脂酰-2-油酰磷脂酰丝氨酸可以诱导含胆固醇的脂质双层发生相分离，并保护胆固醇免受胆固醇氧化酶的侵害。最后，改变的跨双层分布的胆固醇导致flotillin-1重新定位到内吞细胞器。这种探针应该是有用的，在未来的研究池中的细胞质小叶的质膜和细胞器的胆固醇。
Cholesterol is an essential component of metazoan cellular membranes and it helps to maintain the structural integrity and fluidity of the plasma membrane. Here, we developed a cholesterol biosensor, termed D4H, based on the fourth domain of Clostridium perfringens theta-toxin, which recognizes cholesterol in the cytosolic leaflet of the plasma membrane and organelles. The D4H probe disassociates from the plasma membrane upon cholesterol extraction and after perturbations in cellular cholesterol trafficking. When used in combination with a recombinant version of the biosensor, we show that plasmalemmal phosphatidylserine is essential for retaining cholesterol in the cytosolic leaflet of the plasma membrane. In vitro experiments reveal that 1-stearoy-2-oleoyl phosphatidylserine can induce phase separation in cholesterol-containing lipid bilayers and shield cholesterol from cholesterol oxidase. Finally, the altered transbilayer distribution of cholesterol causes flotillin-1 to relocalize to endocytic organelles. This probe should be useful in the future to study pools of cholesterol in the cytosolic leaflet of the plasma membrane and organelles.