IMMUNOLOCALIZATION OF TROPOMODULIN, TROPOMYOSIN AND ACTIN IN SPREAD HUMAN ERYTHROCYTE SKELETONS

The human erythrocyte membrane skeleton consists of a network of short actin filaments cross-linked into a hexagonal network by long, flexib le spectrin molecules. The lengths of the short actin filaments (33+/- nm) at the central junctions are proposed to be stabilized and limited by association with tropomyosin and the tropomyosin-binding protein, tropomodulin. Here, we use immunogold labelling followed by negative s taining to specifically localize tropomodulin, tropomyosin and actin t o the sites of the central junctions in spread membrane skeletons. In addition to negative staining, immunogold labelling for tropomodulin a t the sites of the central junctions was also visualized by a quick-fr eeze, deep-etch, rotary-replication technique, These experiments confi rm previous indirect evidence that the short filaments at the central junctions are indeed actin filaments and provide the first direct evid ence that tropomodulin and tropomyosin are associated with the erythro cyte actin filaments in situ.