Johnson S A, Stinson B M, Go M S, Carmona L M, Reminick J I, Fang X, Baumgart T
Department of Chemistry, University of Pennsylvania, Philadelphia, PA, 19104, USA.
Biochim Biophys Acta. 2010 Jul;1798(7):1427-35. doi: 10.1016/j.bbamem.2010.03.009. Epub 2010 Mar 15.
Liquid-ordered (Lo) and liquid-disordered (Ld) phase coexistence has been suggested to partition the plasma membrane of biological cells into lateral compartments, allowing for enrichment or depletion of functionally relevant molecules. This dynamic partitioning might be involved in fine-tuning cellular signaling fidelity through coupling to the plasma membrane protein and lipid composition. In earlier work, giant plasma membrane vesicles, obtained by chemically induced blebbing from cultured cells, were observed to reversibly phase segregate at temperatures significantly below 37 degrees C. In this contribution, we compare the temperature dependence of fluid phase segregation in HeLa and rat basophilic leukemia (RBL) cells. We find an essentially monotonic temperature dependence of the number of phase-separated vesicles in both cell types. We also observe a strikingly broad distribution of phase transition temperatures in both cell types. The binding of peripheral proteins, such as cholera toxin subunit B (CTB), as well as Annexin V, is observed to modulate phase transition temperatures, indicating that peripheral protein binding may be a regulator for lateral heterogeneity in vivo. The partitioning of numerous signal protein anchors and full length proteins is investigated. We find Lo phase partitioning for several proteins assumed in the literature to be membrane raft associated, but observe deviations from this expectation for other proteins, including caveolin-1.
液态有序（Lo）相和液态无序（Ld）相共存被认为可将生物细胞膜分隔成侧向区室，从而使功能相关分子得以富集或耗竭。这种动态分隔可能通过与质膜蛋白和脂质组成的耦合参与微调细胞信号转导保真度。在早期工作中，通过化学诱导培养细胞形成泡状突起获得的巨大质膜囊泡，在显著低于37摄氏度的温度下被观察到会发生可逆的相分离。在本研究中，我们比较了HeLa细胞和大鼠嗜碱性粒细胞白血病（RBL）细胞中液相分离的温度依赖性。我们发现两种细胞类型中相分离囊泡数量的温度依赖性基本呈单调变化。我们还观察到两种细胞类型的相变温度分布都非常宽泛。观察到外周蛋白如霍乱毒素B亚基（CTB）以及膜联蛋白V的结合会调节相变温度，这表明外周蛋白结合可能是体内侧向异质性的一种调节因子。我们研究了多种信号蛋白锚定物和全长蛋白的分隔情况。我们发现文献中假定与膜筏相关的几种蛋白存在Lo相分隔，但也观察到其他蛋白（包括小窝蛋白-1）偏离了这一预期。
