Department of Medicine, University of California, San Francisco, California, USA.
Biophys J. 2011 Jun 22;100(12):2936-45. doi: 10.1016/j.bpj.2011.05.012.
Tetramers of aquaporin-4 (AQP4) water channels form supramolecular assemblies in cell membranes called orthogonal arrays of particles (OAPs). We previously reported evidence that a short (M23) AQP4 isoform produced by alternative splicing forms OAPs by an intermolecular N-terminus interaction, whereas the full-length (M1) AQP4 isoform does not by itself form OAPs but can coassemble with M23 in OAPs as heterotetramers. Here, we developed a model to predict number distributions of OAP size, shape, and composition as a function M23:M1 molar ratio. Model specifications included: random tetrameric assembly of M1 with M23; intertetramer associations between M23 and M23, but not between M1 and M23 or M1; and a free energy constraint limiting OAP size. Model predictions were tested by total internal reflection fluorescence microscopy of AQP4-green-fluorescent protein chimeras and native gel electrophoresis of cells expressing different M23:M1 ratios. Experimentally validated model predictions included: 1), greatly increased OAP size with increasing M23:M1 ratio; 2), marked heterogeneity in OAP size at fixed M23:M1, with increased M23 fraction in larger OAPs; and 3), preferential M1 localization at the periphery of OAPs. The model was also applied to test predictions about binding to AQP4 OAPs of a pathogenic AQP4 autoantibody found in the neuroinflammatory demyelinating disease neuromyelitis optica. Our model of AQP4 OAPs links a molecular-level interaction of AQP4 with its supramolecular assembly in cell membranes.
水通道蛋白 4（AQP4）四聚体在细胞膜中形成称为正交颗粒排列（OAP）的超分子组装体。我们之前的报告表明，通过选择性剪接产生的短（M23）AQP4 异构体通过分子间的 N 端相互作用形成 OAP，而全长（M1）AQP4 异构体本身不能形成 OAP，但可以与 M23 共同组装成 OAP 作为异四聚体。在这里，我们开发了一个模型来预测 OAP 大小、形状和组成的数量分布作为 M23:M1 摩尔比的函数。模型规范包括：M1 与 M23 的随机四聚体组装；M23 与 M23 之间的 tetramer 之间的关联，但 M1 与 M23 或 M1 之间没有关联；以及限制 OAP 大小的自由能约束。通过 AQP4-绿色荧光蛋白嵌合体的全内反射荧光显微镜和表达不同 M23:M1 比例的细胞的天然凝胶电泳测试了模型预测。经过实验验证的模型预测包括：1），随着 M23:M1 比值的增加，OAP 尺寸大大增加；2），在固定的 M23:M1 下，OAP 尺寸的显着异质性，较大的 OAP 中增加了 M23 分数；3），M1 优先定位于 OAP 的外围。该模型还应用于测试在神经炎症性脱髓鞘疾病视神经脊髓炎中发现的致病性 AQP4 自身抗体与 AQP4 OAP 结合的预测。我们的 AQP4 OAP 模型将 AQP4 与其在细胞膜中的超分子组装之间的分子水平相互作用联系起来。
