Temperature-sensitive non-fusing myoblast variant and spontaneous revertant: isolation and characterization

A stable, temperature-sensitive, non-fusing variant of the L6 rat myoblast cell line has been isolated following mild EMS-induced mutagenesis. At the permissive temperature (37 degrees C), the growth characteristics and developmental pattern of the tsA1 variant are essentially identical to those of the parental L6D0 line at either 37 degrees C or 40 degrees C. At the nonpermissive temperature (40 degrees C), the tsA1 variant grows normally but does not align, fuse, or synthesize detectable amounts of beta-tropomyosin or myosin LC2. A peptide corresponding to myosin LClemb is barely detectable. The temperature-sensitive period spans the interval from 4 to 72 h post-plating with a midpoint at approximately 40 h. Under standard culture conditions, commitment to terminal differentiation occurs between days 3 and 4, and alignment and fusion begin on days 4 and 5, respectively. Thus, the temperature-sensitive event occurs very early in the L6 developmental program. A spontaneous revertant of the temperature-sensitive phenotype (tsA1 [R3]) exhibits recovery of the capacities to align, fuse, and synthesize the repertoire of muscle-specific proteins, suggesting that a single pleiotropic mutation in the tsA1 variant may regulate several stages in L6 myogenesis.