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 ectopic expression of daughterless alone, as indicated by exit from the cell cycle, syncytia formation, and the presence of muscle myosin fibrils. Myogenic conversion can be potentiated by the coexpression of DMEF2 and nautilus with daughterless. In RT-PCR assays Schneider cells express two mesodermal markers, 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 encode immunoprecipitable proteins. We used RNA-i to demonstrate that both endogenous nautilus expression and DMEF2 expression are required for the myogenic conversion of Schneider cells by daughterless. Coexpression of twist blocks conversion by daughterless but twist dsRNA has no effect. Our results indicate that Schneider cells are of mesodermal origin and that myogenic conversion with ectopic expression of daughterless occurs by raising the levels of daughterless protein sufficiently to allow the formation of nautilus/daughterless heterodimers. The effectiveness of RNA-i is dependent upon protein 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 amenable to the application of RNA-i. These results support the conclusion that nautilus is a myogenic factor in Drosophila tissue culture cells with a functional role similar to that of vertebrate MyoD. This is discussed with regard to the in vivo functions of nautilus.
Wei
Q
Q
Laboratory of Biochemistry, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Marchler
G
G
Edington
K
K
Karsch-Mizrachi
I
I
Paterson
B M
BM
D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins; D015415:Biomarkers; D002454:Cell Differentiation; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D029721:Drosophila Proteins; D004331:Drosophila melanogaster; D005786:Gene Expression Regulation; D007301:Insect Hormones / Q000502:physiology*; D019476:Insect Proteins / Q000235:genetics / Q000378:metabolism; D064326:MEF2 Transcription Factors; D008648:Mesoderm / Q000166:cytology* / Q000502:physiology; D009132:Muscles / Q000166:cytology* / Q000502:physiology; D018006:Myogenic Regulatory Factors; D009687:Nuclear Proteins / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism / Q000502:physiology*; D014158:Transcription, Genetic; D051879:Twist-Related Protein 1
D051792:Basic Helix-Loop-Helix Proteins; D015415:Biomarkers; D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D007301:Insect Hormones; D019476:Insect Proteins; D064326:MEF2 Transcription Factors; D018006:Myogenic Regulatory Factors; D009687:Nuclear Proteins; D012333:RNA, Messenger; D014157:Transcription Factors; D051879:Twist-Related Protein 1; C090080:Da protein, Drosophila; C499409:Mef2 protein, Drosophila; C497753:Twi protein, Drosophila