Comparison of several promoters and polyadenylation signals for use in heterologous gene expression in cultured Drosophila cells.
We have directly compared the ability of four promoters and three polyadenylation (poly(A)) signals to direct heterologous gene expression in stably transfected Drosophila melanogaster S2 cells. We compared two constitutive Drosophila promoters, the actin 5C distal promoter and the alpha 1-tubulin promoter, with the tightly regulated Drosophila metallothionein (Mtn) promoter and the Bombyx mori fibroin promoter. We find that the actin 5C and induced Mtn promoters generate comparable high levels of RNA and protein in this system. The alpha 1-tubulin promoter generates about four-fold lower levels, and the fibroin promoter shows no detectable activity in S2 cells. Interestingly, genes expressed from the constitutive actin 5C and alpha 1-tubulin promoters are consistently present at three- to four-fold lower copy numbers than genes expressed from the inducible Mtn promoter or the inactive fibroin promoter. Poly(A) signals of both mammalian (SV40) and Drosophila (Mtn) origin efficiently directed stable RNA synthesis in S2 cells, and, as in mammalian cells, the SV40 late poly(A) signal was more efficient than the SV40 early poly(A) signal. Thus the process of polyadenylation appears to be conserved between mammalian and Drosophila cells.
Angelichio
M L
ML
Department of Gene Expression Sciences, SmithKline Beecham Pharmaceuticals, King of Prussia, PA 19406.
Beck
J A
JA
Johansen
H
H
Ivey-Hoyle
M
M
D000199:Actins / Q000235:genetics; D000818:Animals; D015153:Blotting, Western; D002478:Cells, Cultured; D004331:Drosophila melanogaster / Q000235:genetics*; D005349:Fibroins / Q000235:genetics; D015870:Gene Expression; D005822:Genetic Vectors; D015699:HIV Envelope Protein gp120 / Q000235:genetics; D008668:Metallothionein / Q000235:genetics; D010957:Plasmids; D011061:Poly A / Q000235:genetics / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D013539:Simian virus 40 / Q000235:genetics; D010959:Tissue Plasminogen Activator / Q000235:genetics; D014162:Transfection; D014404:Tubulin / Q000235:genetics
D000199:Actins; D015699:HIV Envelope Protein gp120; D014404:Tubulin; D011061:Poly A; D005349:Fibroins; D008668:Metallothionein; D010959:Tissue Plasminogen Activator