Journal
MICROBIOLOGY-SGM
Volume 146, Issue -, Pages 2665-2670Publisher
MICROBIOLOGY SOC
DOI: 10.1099/00221287-146-10-2665
Keywords
peptide nucleic acid; bacteria; lipopolysaccharide
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Antisense peptide nucleic acids (PNAs) can inhibit Escherichia coli gene expression and cell growth through sequence-specific RNA binding, and this opens possibilities for novel anti-infective agents and tools for microbial functional genomics. However, the cellular effects of PNAs are limited relative to effects in cell extracts, presumably because of cell barrier components such as the outer-membrane lipopolysaccharide (LPS) layer or drug efflux pumps, both of which function to exclude antibiotics and other foreign molecules. To evaluate the importance of such cellular factors on RNA effects, the authors developed a positive assay for antisense inhibition by targeting the lac operon repressor and compared RNA susceptibilities in mutant and wild-type E. coli by assessing lacZ induction. Strains with defective LPS (AS19 and D22) were more permeable to the antibiotic nitrocefin and more susceptible to RNA than the wild-type. Also, RNA potency was improved in wild-type cells grown in the presence of certain cell-wall-permeabilizing agents. In contrast, the activities of the Acr and Emr drug efflux pumps were not found to affect RNA susceptibility. The results show that the LPS layer is a major barrier against cell entry, but PNAs that can enter E. coli are likely to remain active inside cells.
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