Abstract
We address the controversy of whether mature myofibrils can form in the presence of taxol, a microtubule‐stabilizing compound. Previous electron microscopic studies reported the absence of actin filaments and Z‐bands in taxol‐treated myocytes [Antin et al., 1981: J Cell Biol 90:300–308; Toyoma et al., 1982: Proc Natl Acad Sci USA 79:6556–6560]. Quail skeletal myoblasts were isolated from 10‐day‐old embryos and grown in the presence or absence of taxol. Taxol inhibited the formation of multinucleated elongated myotubes. Myocytes cultured in the continual presence of taxol progressed from rounded to stellate shapes. Groups of myocytes that were clustered together after the isolation procedure fused in the presence of taxol but did not form elongated myotubes. Actin filaments and actin‐binding proteins were detected with several different fluorescent probes in all myofibrils that formed in the presence of taxol. The Z‐bands contained both alpha‐actinin and titin, and the typical arrays of A‐Bands were always associated with actin filaments in the myofibrils. Myofibril formation was followed by fixing cells each day in culture and staining with probes for actin, muscle‐specific alpha‐actinin, myosin II, nebulin, troponin, tropomyosin, and non‐muscle myosin II. Small linear aggregates of alpha‐actinin or Z‐bodies, premyofibrils, were detected at the edges of the myocytes and in the arms of the taxol‐treated cells and were always associated with actin filaments. Non‐muscle myosin II was detected at the edges of the taxol‐treated cells. Removal of the taxol drug led to the cells assuming a normal compact elongated shape. During the recovery process, additional myofibrils formed at the spreading edges of these elongated and thicker myotubes. Staining of these taxol‐recovering cells with specific fluorescent reagents reveals three different classes of actin fibers. These results are consistent with a model of myofibrillogenesis that involves the transition of premyofibrils to mature myofibrils. Cell Motil. Cytoskeleton 58:39–52, 2004. © 2004 Wiley‐Liss, Inc.