The distribution of Na+,K(+)-ATPase and 5A11 antigen in apical microvilli of the retinal pigment epithelium is unrelated to alpha-spectrin.
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.
Rizzolo
L J
LJ
Department of Surgery, Yale University School of Medicine, New Haven, CT 06520-8062, USA.
Zhou
S
S
D000818:Animals; D017487:Ankyrins / Q000032:analysis; D000954:Antigens, Surface / Q000032:analysis*; D002454:Cell Differentiation / Q000502:physiology; D002462:Cell Membrane / Q000737:chemistry / Q000201:enzymology; D016764:Cell Polarity / Q000502:physiology; D002642:Chick Embryo; D003599:Cytoskeleton / Q000502:physiology; D005455:Fluorescent Antibody Technique; D007536:Isomerism; D008871:Microvilli / Q000737:chemistry* / Q000201:enzymology; D017830:Octoxynol; D010857:Pigment Epithelium of Eye / Q000737:chemistry* / Q000166:cytology / Q000201:enzymology; D000254:Sodium-Potassium-Exchanging ATPase / Q000032:analysis*; D013049:Spectrin / Q000032:analysis*; D013997:Time Factors
D017487:Ankyrins; D000954:Antigens, Surface; D013049:Spectrin; D017830:Octoxynol; D000254:Sodium-Potassium-Exchanging ATPase