Actin isoform compartments in chicken gizzard smooth muscle cells

Abstract

Differentiated smooth muscle cells typically contain a mixture of muscle (alpha and gamma) and cytoplasmic (beta and gamma) actin isoforms. Of the cytoplasmic actins the beta-isoform is the more dominant, making up from 10% to 30% of the total actin complement. Employing an antibody raised against the N-terminal peptide specific to beta-actin, which labels only the beta-isoform on two-dimensional gel immunoblots, we have shown that this isoform has a restricted localisation in smooth muscle. Using double-label immunofluorescence and immunoelectron microscopy of ultrathin sections of chicken gizzard, beta-actin was localised in the dense bodies and in longitudinal channels linking consecutive dense bodies that were also occupied by desmin. It was additionally found in the membrane-associated dense plaques, but was excluded from the actomyosin-containing regions of the contractile apparatus. Taken together with earlier results these findings identify a cytoskeletal compartment containing intermediate filaments, cytoplasmic actin and the actin cross-linking protein filamin. Using an antibody specific only for muscle actin, labelling was found generally around the myosin filaments of the contractile apparatus, but was absent from the core of the dense bodies that contained beta-actin. Thus, if dense bodies act as dual-purpose anchorage sites, for the cytoskeletal actin and the contractile actin, the thin filaments of the contractile apparatus must be anchored at the periphery of the dense bodies. A model of the structural organisation of the cell is presented and the possible roles of the cytoskeleton are discussed.