RNA interference demonstrates a role for nautilus in the myogenic conversion of Schneider cells by daughterless

Schneider SL2 cells activate the myogenic program in response to the ectopi c expression of daughterless alone, as indicated by exit from the cell cycl e, syncytia formation, and the presence of muscle myosin fibrils. Myogenic conversion can be potentiated by the coexpression of DMEF2 and nautilus wit h daughterless. In RT-PCR assays Schneider cells express two mesodermal mar kers, nautilus and DMEF2 mRNAs, as well as very low levels of daughterless mRNA but no twist. Full-length RT-PCR products for nautilus and DMEF2 encod e immunoprecipitable proteins. We used RNA-i to demonstrate that both endog enous nautilus expression and DMEF2 expression are required for the myogeni c conversion of Schneider cells by daughterless. Coexpression of twist bloc ks conversion by daughterless but twist dsRNA has no effect. Our results in dicate that Schneider cells are of mesodermal origin and that myogenic conv ersion with ectopic expression of daughterless occurs by raising the levels of daughterless protein sufficiently to allow the formation of nautilus/ d aughterless heterodimers. The effectiveness of RNA-i is dependent upon prot ein half-life. Genes encoding proteins with relatively short half-lives (10 h), such as nautilus or HSF, are efficiently silenced, whereas more stable proteins, such as cytoplasmic actin or beta -galactosidase, are less amena ble to the application of RNA-i. These results support the conclusion that nautilus is a myogenic factor in Drosophila tissue culture cells with a fun ctional role similar to that of vertebrate MyoD. This is discussed with reg ard to the in vivo functions of nautilus. (C) 2000 Academic Press.