Induction of dystrophin localization in cultured Xenopus muscle cells by latex beads

Abstract

The distribution of dystrophin in Xenopus myotomal muscle cells was examined in conventional and confocal immunofluorescence microscopy. By labeling dissociated single muscle fibers with a monoclonal or a polyclonal antibody against dystrophin, we found that dystrophin is ten times more concentrated at the myotendinous junction (MTJ) than at the extrajunctional sarcolemma. At the MTJ, dystrophin lines the membrane invaginations where myofibrils attach to the membrane. It is colocalized with talin, but is not related to the distribution of acetylcholine receptors (AChRs) which are clustered at the postsynaptic membrane in the vicinity of the MTJ in these fibers. We found that the localization of dystrophin can be induced in cultured Xenopus myotomal muscle cells by treating them with polystyrene latex beads. Dystrophin is discretely localized at the bead-muscle contacts. With electron microscopy, a sarcolemma specialization with all the salient features of the MTJ, including basal lamina-lined membrane invaginations along which myofibrils make attachment. Although these beads also induce clustering of AChRs, the patterns of dystrophin and AChR localization are distinct. The appearance of dystrophin at the bead-contacted sarcolemma is coincident with the development of the membrane invaginations. This, together with its concentration along membrane invaginations at the MTJ in vivo, suggests a role for dystrophin in the formation of this junctional specialization. Since the signal for MTJ development can be presented to cultured muscle cells in a temporally and spatially controlled manner by beads, this system offers a simple model for analyzing the mechanism of this sarcolemma specialization.