Lin Wan-Chen, Blanchette Craig D, Ratto Timothy V, Longo Marjorie L
Biophysics Graduate Group, Division of Biological Sciences, University of California, Davis, California 95616, USA.
Biophys J. 2006 Jan 1;90(1):228-37. doi: 10.1529/biophysj.105.067066. Epub 2005 Oct 7.
A fundamental attribute of cell membranes is transmembrane asymmetry, specifically the formation of ordered phase domains in one leaflet that are compositionally different from the opposing leaflet of the bilayer. Using model membrane systems, many previous studies have demonstrated the formation of ordered phase domains that display complete transmembrane symmetry; but there have been few reports on the more biologically relevant asymmetric membrane structures. Here we report on a combined atomic force microscopy and fluorescence microscopy study whereby we observe three different states of transmembrane symmetry in phase-separated supported lipid bilayers formed by vesicle fusion. We find that if the leaflets differ in gel-phase area fraction, then the smaller domains in one leaflet are in registry with the larger domains in the other leaflet and the system is dynamic. In a presumed lipid flip-flop process similar to Ostwald ripening, the smaller domains in one leaflet erode away whereas the large domains in the other leaflet grow until complete compositional asymmetry is reached and remains stable. We have quantified this evolution and determined that the lipid flip-flop event happens most frequently at the interface between symmetric and asymmetric DSPC domains. If both leaflets have identical area fraction of gel-phase, gel-phase domains are in registry and are static in comparison to the first state. The stability of these three DSPC domain distributions, the degree of registry observed, and the domain immobility have biological significance with regards to maintenance of lipid asymmetry in living cell membranes, communication between inner leaflet and outer leaflet, membrane adhesion, and raft mobility.
细胞膜的一个基本特性是跨膜不对称性，具体表现为在一个小叶中形成有序相域，其组成与双层膜的相对小叶不同。使用模型膜系统，许多先前的研究已经证明了形成具有完全跨膜对称性的有序相域；但关于更具生物学相关性的不对称膜结构的报道却很少。在这里，我们报告了一项结合原子力显微镜和荧光显微镜的研究，通过该研究我们观察了由囊泡融合形成的相分离支撑脂质双层中的三种不同跨膜对称状态。我们发现，如果小叶在凝胶相面积分数上不同，那么一个小叶中的较小域与另一个小叶中的较大域对齐，并且系统是动态的。在一个类似于奥斯特瓦尔德熟化的假定脂质翻转过程中，一个小叶中的较小域逐渐侵蚀消失，而另一个小叶中的大域生长，直到达到完全的组成不对称并保持稳定。我们已经对这种演变进行了量化，并确定脂质翻转事件最常发生在对称和不对称二硬脂酰磷脂酰胆碱（DSPC）域之间的界面处。如果两个小叶具有相同的凝胶相面积分数，凝胶相域会对齐，并且与第一种状态相比是静态的。这三种DSPC域分布的稳定性、观察到的对齐程度以及域的固定性对于维持活细胞膜中的脂质不对称性、内膜小叶和外膜小叶之间的通信、膜粘附以及筏的流动性具有生物学意义。
