LOCAL ACCUMULATION OF ALPHA-SPECTRIN-RELATED PROTEIN UNDER PLASMA-MEMBRANE DURING CAPPING AND PHAGOCYTOSIS IN ACANTHAMOEBA

During capping and phagocytosis the interaction between clustered cell surface receptors and the submembraneous actin-based skeleton may be mediated by spectrin-like proteins. To test this possibility we examin ed the localization of an a-spectrin immunoanalogue, that had been pre viously identified in whole extracts of Acanthamoeba, during capping o f Con A receptors and during phagocytosis of Con A-coated yeast. Durin g capping alpha-spectrin and filamentous actin co-migrated with the Co n A receptors and accumulated in the region of cap formation, as demon strated by double immunofluorescence studies. Immunoelectron microscop y revealed submembraneous location of alpha-spectrin in cells exposed to Con A, both at the time of initial cross-linking and during accumul ation of alpha-spectrin in the region of the cap. Phagocytosis studies showed that alpha-spectrin and actin filaments were concentrated arou nd phagocytic cups that enclosed Con A-coated yeast upon internalizati on. The proteins also surrounded nascent phagosomes present in the vic inity of the plasma membrane but were absent at the later time point o f phagosome maturation. These data demonstrate a correlation between c lustering of cell surface receptors and submembraneous localization of alpha-spectrin, suggesting an involvement of spectrin-like proteins i n mediating the interaction of receptor clusters with the actin cytosk eleton. (C) 1997 Wiley-Liss, Inc.