Rizzolo L J, Zhou S
Department of Surgery, Yale University School of Medicine, New Haven, CT 06520-8062, USA.
J Cell Sci. 1995 Nov;108 ( Pt 11):3623-33. doi: 10.1242/jcs.108.11.3623.
The retinal pigment epithelium was used to study the relationship between the cortical cytoskeleton and two plasma membrane proteins that associate with it. These proteins were the Na+,K(+)-ATPase, an ion pump, and the 5A11 antigen, a member of the immunoglobulin superfamily of receptor proteins. The cytoskeleton was marked by two of its constituents, alpha-spectrin and ankyrin. Ankyrin links the Na+,K(+)-ATPase to spectrin in many cells. The RPE is of interest, because unlike most epithelia it distributes the Na+,K(+)-ATPase to the apical membrane. The development of polarity was studied during chick embryogenesis. On embryonic day 6 (E6), each of these proteins was observed in the apical and lateral plasma membranes. As development proceeded, only the Na+,K(+)-ATPase was removed from the lateral membranes. Beginning on E12, ankyrin, spectrin and 5A11 appeared together in patches along the basal plasma membrane. By E16, these patches coalesced into a uniform distribution along the basal membrane. At the apical pole, alpha-spectrin appeared near the base of the microvilli, but was undetected in the microvilli themselves. This distribution resembled the distribution of alpha-spectrin in the intestine and proximal kidney tubule. By contrast, a pool of ankyrin and 5A11 and nearly all the Na+,K(+)-ATPase appeared in the microvilli. Despite its segregation from alpha-spectrin, the Na+,K(+)-ATPase appeared to associate with a macromolecular complex, as judged by extraction with Triton X-100. Changes in spectrin distribution could not be related to changes in isoform expression, as only one isoform of beta-spectrin was detected by co-immunoprecipitation with alpha-spectrin. By contrast, multiple ankyrin-like peptides could be identified by immunoblotting. These data illustrate some of the unique properties of RPE microvilli. These properties prevent the Na+,K(+)-ATPase from complexing with the alpha-spectrin-based cytoskeleton by sequestering the enzyme into the compartment where its activity is required.
视网膜色素上皮细胞被用于研究皮质细胞骨架与两种与其相关的质膜蛋白之间的关系。这两种蛋白分别是离子泵钠钾ATP酶和受体蛋白免疫球蛋白超家族成员5A11抗原。细胞骨架由其两种成分α-血影蛋白和锚蛋白标记。在许多细胞中，锚蛋白将钠钾ATP酶与血影蛋白相连。视网膜色素上皮细胞很受关注，因为与大多数上皮细胞不同，它将钠钾ATP酶分布于顶端膜。在鸡胚胎发育过程中研究了极性的形成。在胚胎第6天（E6），这些蛋白中的每一种都在顶端和侧面质膜中被观察到。随着发育的进行，只有钠钾ATP酶从侧面膜上移除。从E12开始，锚蛋白、血影蛋白和5A11一起出现在沿基底质膜的斑块中。到E16时，这些斑块合并成沿基底膜的均匀分布。在顶端极，α-血影蛋白出现在微绒毛基部附近，但在微绒毛本身中未检测到。这种分布类似于α-血影蛋白在肠道和近端肾小管中的分布。相比之下，一小部分锚蛋白和5A11以及几乎所有的钠钾ATP酶出现在微绒毛中。尽管钠钾ATP酶与α-血影蛋白分离，但通过用Triton X-100提取判断，它似乎与一个大分子复合物相关。血影蛋白分布的变化与同工型表达的变化无关，因为通过与α-血影蛋白共免疫沉淀仅检测到一种β-血影蛋白同工型。相比之下，通过免疫印迹可鉴定出多种锚蛋白样肽。这些数据说明了视网膜色素上皮细胞微绒毛的一些独特特性。这些特性通过将该酶隔离到需要其活性的区室中，阻止钠钾ATP酶与基于α-血影蛋白的细胞骨架形成复合物。
