Gene switching in myogenesis: differential expression of the chicken actin multigene family.

We described the construction of an alpha-actin complementary deoxyribonucleic acid (cDNA) clone, pAC269 [Schwartz, R. J., Haron, J. A., Rothblum, K. N., & Dugaiczyk, A. (1980) Biochemistry 19, 5883], that was used as a hybridization probe in the current investigation to examine the induction of actin messenger ribonucleic acid (mRNA) during myogenesis. A Tm difference of 10-13 degrees C between skeletal muscle alpha-actin and nonmuscle beta- and gamma-actin mRNAs and pAC269 allowed us to establish the highly stringent hybridization conditions necessary to measure separately the content of alpha-actin mRNA and beta- and gamma-actin mRNA during muscle development in culture. We observed low levels of alpha-actin mRNA (approximately 130 molecules/cell) in replicating prefusion myoblasts. The vast majority of actin mRNA (2000 molecules/cell) present at this stage was accounted for by beta- and gamma-actin mRNA. Beginning at myoblast fusion, alpha-actin mRNA accumulated and within 30 h reached a level 270-fold greater than that observed in the undifferentiated state. At 95 h in culture when myotube formation was completed, alpha-actin content was at its peak (36 000 molecules/nucleus). Conversely, beta- and gamma-actin mRNA content began to decline at the beginning of fusion, and by the end of myotube formation beta- and gamma-actin mRNAs were undetectable by our techniques. A rapid depression of alpha-actin mRNA levels was observed after 95 h in the absence of cell death. At 6 days after the initiation of myotube formation, the content of alpha-actin mRNA was reduced by 80% in comparison of peak values and remained at that level. The switching of actin mRNA species was inhibited in myoblasts treated with bdU. The accumulation of alpha-actin mRNA and the disappearance of beta- and gamma-actin mRNA were observed following the reversal of the bdU block and coincident with the onset of myoblast fusion. We found that the expression of actin genes within the actin multigene family is switched in myogenesis through a strict developmental pattern.