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Inhibition of gene expression in Escherichia coli by antisense phosphorodiamidate morpholino oligomers.
|Title||Inhibition of gene expression in Escherichia coli by antisense phosphorodiamidate morpholino oligomers.|
|Publication Type||Journal Article|
|Year of Publication||2003|
|Authors||Geller BL, Deere JD, Stein DA, Kroeker AD, Moulton HM, Iversen PL|
|Journal||Antimicrobial agents and chemotherapy|
|Date Published||2003 Oct|
|Keywords||Acyl Carrier Protein, Amino Acid Sequence, Bacterial Proteins, Base Sequence, beta-Galactosidase, Cell Membrane Permeability, Dose-Response Relationship, Drug, Escherichia coli, Escherichia coli Proteins, Gene Expression, Genes, myc, Genes, Reporter, Lac Repressors, Luciferases, Membrane Transport Proteins, Morpholines, Morpholinos, Oligonucleotides, Antisense, Repressor Proteins, RNA, Ribosomal, 16S|
Antisense phosphorodiamidate morpholino oligomers (PMOs) were tested for the ability to inhibit gene expression in Escherichia coli. PMOs targeted to either a myc-luciferase reporter gene product or 16S rRNA did not inhibit luciferase expression or growth. However, in a strain with defective lipopolysaccharide (lpxA mutant), which has a leaky outer membrane, PMOs targeted to the myc-luciferase or acyl carrier protein (acpP) mRNA significantly inhibited their targets in a dose-dependent response. A significant improvement was made by covalently joining the peptide (KFF)(3)KC to the end of PMOs. In strains with an intact outer membrane, (KFF)(3)KC-myc PMO inhibited luciferase expression by 63%. A second (KFF)(3)KC-PMO conjugate targeted to lacI mRNA induced beta-galactosidase in a dose-dependent response. The end of the PMO to which (KFF)(3)KC is attached affected the efficiency of target inhibition but in various ways depending on the PMO. Another peptide-lacI PMO conjugate was synthesized with the cationic peptide CRRRQRRKKR and was found not to induce beta-galactosidase. We conclude that the outer membrane of E. coli inhibits entry of PMOs and that (KFF)(3)KC-PMO conjugates are transported across both membranes and specifically inhibit expression of their genetic targets.
|Alternate Journal||Antimicrob. Agents Chemother.|