Biochemical Characterization of Ezrin-Actin Interaction (*)

Abstract

The highly related actin isoforms are thought to have different functions. We recently demonstrated a polarized distribution of actin isoforms in gastric parietal cells and association of gastric ezrin with the cytoplasmic β-actin isoform (Yao, X., Chaponnier, C., Gabbiani, G., and Forte, J. G.(1995) Mol. Biol. Cell. 6, 541-557). Here we used ultrastructural immunocytochemistry to verify that β-actin is located within canalicular microvilli and the apical cortex of parietal cells, similar to the localization reported for ezrin. Furthermore, we tested whether ezrin binds preferentially to cytoplasmic β-actin compared with the skeletal muscle α-actin isoform. Purified cytoplasmic β-actin (from erythrocytes) and skeletal α-actin were assembled with gastric ezrin. Co-sedimentation experiments showed that gastric ezrin selectively co-pelleted with the β-actin isoform and only very poorly with α-actin. Binding of erythrocytic β-actin to ezrin is saturable with a molar ratio of 1:10 (ezrin:actin) and a dissociation constant 4.6 × 10M. In addition, ezrin promoted pyrene-labeled actin assembly, with predominant effects on filament elongation and a distinct preference for β-actin compared with α-actin. Given these isoform-selective associations, we speculate that actin isoforms might segregate into different functional domains and exert specificity by interacting with isoform-orientated binding proteins.