The vinculin/sarcomeric-alpha-actinin/alpha-actin nexus in cultured cardiac myocytes

Abstract

Abstract. Experiments are described supporting the proposition that the assembly of stress fibers in nonmuscle cells and the assembly of myofibrils in cardiac cells share conserved mechanisms. Double staining with a battery of labeled antibodies against membraneassociated proteins, myofibrillar proteins, and stress fiber proteins reveals the following: (a) dissociated, cultured cardiac myocytes reconstitute intercalated discs consisting of adherens junctions (AJs) and desmosomes at sites of cell-cell contact and sub-sarcolemmal adhesion plaques (SAPs) at sites of cellsubstrate contact; (b) each AJ or SAP associates proximally with a striated myofibril, and conversely every striated myofibril is capped at either end by an AJ or a SAP; (C) the invariant association between a given myofibril and its SAP is especially prominent at the earliest stages of myofibrillogenesis; nascent myofibrils are capped by oppositely oriented SAPs; (d) the insertion of nascent myofibrils into AJs or into SAPs invariably involves vinculin, a-actin, and sarcomeric ot-actinin (s-ot-actinin); (e) AJs are positive for A-CAM but negative for talin and integrin; SAPs lack A-CAM but are positive for talin and integrin; (f) in cardiac cells all ot-actinin-containing structures invariably are positive for the sarcomeric isoform, o~-actin and related sarcomeric proteins; they lack non-sot-actinin, -y-actin, and caldesmon; (g) in fibroblasts all ot-actinin-containing structures are positive for the non-sarcomeric isoform, -y-actin, and related nonsarcomeric proteins, including caldesmon; and (h) myocytes differ from all other types of adherent cultured cells in that they do not assemble authentic stress fibers; instead they assemble stress fiber-like structures of linearly aligned I-Z-I-like complexes consisting exclusively of sarcomeric proteins. CONSIDERABLE information has accumulated regarding the molecular composition and possible functions of the submembranous complexes into which stress fibers insert (4,5,11,26,34,46). In cell-cell junctions of the adherens type the distal tips of the microfilament bundles insert into submembranous F-actin/o~-actinin/vinculin complexes that often involve additional membrane-associated molecules such as A-CAM (31, 60), radixin (58) etc. A somewhat different class of submembranous complexes characterizes the termini of stress fibers where they insert into the cell-substrate junctions of spread, nontranslocating cultured ceils. These F-actin/ot-actinin/vinculin complexes, known as adhesion plaques (APs),t are also positive for talin (11), paxillin (59), integrin (17), and other proteins. APs are also sites for a Ca2 § protease (5) and a1. Abbreviations used in this paper: AJ, adherens junction; AP, adhesion plaque; MHC, myosin heavy chain; s, sarcomeric; SAP, subsarcolemmal adhesion plaque.protein kinase C (35) suggesting they may be involved in transmembrane signaling. In both the adherens junction (AJs) and the APs, ct-actinin appears to play a bifunctional role. Distally, facing the inner surface of the c...

Cytoskeletal domains in the activated platelet

“…For example, adhesion plaques and stress fibers contain alpha-actinin [24], while microvilli contain villin and fimbrin [22,23]. In the sarcomere of striated muscle, tropomyosin is distributed along the thin filaments, while gelsolin and CapZ are located at the Z-band, and myosin II forms the thick filaments that overlap but are not superimposable upon the actin [26].…”

Differentiation, extracellular matrix synthesis, and integrin assembly by Drosophila embryo cells cultured on vitronectin and laminin substrates

“…Before sarcomere formation, accumulations of integrins and actin were notable at the contact sites of myotubes with epithelial cells in cultures grown on a laminin substrate. These myoepithelial junctions are likely to act as nucleation sites for the formation of the muscle cytoskeleton, as has been suggested for cell-cell contacts in vertebrate myocytes (Lu et al, 1992).…”

“…Several observations in cultured vertebrate cells indicate that the association of integrins with Z-bands may be a late step in sarcomere formation (Bozyczko et al, 1989;Hilenski et al, 1991) or may not occur at all (Bozyczko et al, 1989;Lu et al, 1992). This would suggest that linkage of a vertebrate cytoskeleton with the extracellular matrix through integrins at Z-bands may be a more stabilizing than initiating event of myogenesis.…”

“…For comparison, during vertebrate myogenesis integrins become concentrated at myotendinous junctions (Bozyczko et al, 1989;Swasdison and Mayne, 1989), and, to varying extents, depending on type of muscle and state of development, are also found at Z bands and at other sites (Bozyczko et al, 1989;Terracio et al, 1989;Lakonishok et al, 1992;Lu et al, 1992). The cytoskeleton may also be attached at Z-bands in other ways.…”

Molecular mechanisms of mechanosensing in muscle development

“…1). ␣-Actinin, the most prominent Z-disk component, initially appears in small aggregates at the sarcolemma during sarcomere assembly (Lu et al, 1992;Dabiri et al, 1997). ␣-Actinin could be recruited to these aggregates by direct interaction with integrin and vinculin, or indirectly through Zasp (Otey et al, 1990;Bois et al, 2005;Kelly and Taylor, 2005;Jani and Schöck, 2007).…”