title: Eukaryotic promoters can direct protein synthesis in Gram-negative bacteria.
abstract: Intracellular bacteria can act as DNA delivery vectors into mammalian cells. Transfer of genetic information can be monitored by screening for cellular expression of a reporter gene under the control of an eukaryotic promoter. However, intracellular bacteria can also efficiently deliver heterologous proteins in the cell cytosol. We have studied the activity of the eukaryotic PCMV promoter in Escherichia coli and Salmonella typhimurium using the lacZ and gfp genes as reporters and determined its strength relative to those of PRSV and PSV40 in E. coli. We found substantial heterologous activity of fragments carrying the PCMV and PRSV promoters, the strength of PRSV being greater than that of PCMV, but did not detect any PSV40 activity in E. coli. The green fluorescent protein (GFP) synthesized in E. coli was transferred to COS-1 cells where it was detectable and stable. Insertion of a transcription terminator or deletion of the bacterial ribosome binding site downstream from PCMV led to the silencing of the promoter in bacteria but not in mammalian cells. These observations should incite to exert caution when interpreting data on the DNA transfer from bacteria to mammallian cells when the genes of interest are under the control of the PCMV or PRSV promoter.
authors.affiliation: Unité des Agents Antibactériens, Institut Pasteur, Paris, France. sgouss@pasteur.fr
authors.forename: Sylvie
authors.initials: S
authors.lastname: Goussard
authors.forename: Catherine
authors.initials: C
authors.lastname: Grillot-Courvalin
authors.forename: Patrice
authors.initials: P
authors.lastname: Courvalin
mesh_terms: D000818:Animals; D019556:COS Cells; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D017930:Genes, Reporter; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D007763:Lac Operon; D008164:Luminescent Proteins / Q000096:biosynthesis*; D010957:Plasmids; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis*; D012486:Salmonella typhimurium / Q000235:genetics* / Q000378:metabolism; D001616:beta-Galactosidase / Q000096:biosynthesis*
chemical_list: D008164:Luminescent Proteins; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins; D015500:Chloramphenicol O-Acetyltransferase; D001616:beta-Galactosidase