RED-CELL MEMBRANES OF ANKYRIN-DEFICIENT NB NB MICE LACK BAND-3 TETRAMERS BUT CONTAIN NORMAL MEMBRANE SKELETONS/

The role of ankyrin in the formation and stabilization of the spectrin -based skeletal meshwork and of band 3 oligomers was studied by charac terizing, in nb/nb mouse red cells, the effect of ankyrin deficiency o n skeletal ultrastructure, band 3-skeleton associations, and band 3 ol igomeric states. Despite severe ankyrin deficiency, nb/nb mouse red ce ll skeletal components formed a relatively uniform two-dimensional hex agonal array of junctional complexes cross-linked by spectrin tetramer s. Treatment of nb/nb ghosts with the nonionic detergent C12E8 (octaet hylene glycol n-dodecyl monoether) resulted in nearly complete extract ion of band 3. The extracted band 3 was present exclusively as band 3 dimers. Fluorescence photobleaching recovery and polarized fluorescenc e depletion measurements showed increases in the laterally (33% vs 10% ) and rotationally (90% vs 76%) mobile fractions of band 3 in Intact n b/nb compared to control red cells. The rotational correlation time of the major fraction of band 3 molecules was 10-fold shorter in nb/nb c ompared to control red cells, indicating a significant relaxation of r otational constraints in nb/nb cells. These data suggest that, althoug h ankyrin plays a major role in strengthening the attachment of the sk eleton to the membrane bilayer, ankyrin is not required for the format ion of a stable two-dimensional spectrin-based skeleton, The absence o f band 3 tetramers in the membrane of ankyrin-deficient red cells sugg ests that ankyrin is required for the formation of stable band 3 tetra mers.