Genetic Evidence for Inhibition of Bacterial Division Protein FtsZ by Berberine

Liam Good,Jem Stach,Jaroslaw M. Boberek

doi:10.1371/journal.pone.0013745

Abstract

BackgroundBerberine is a plant alkaloid that is widely used as an anti-infective in traditional medicine. Escherichia coli exposed to berberine form filaments, suggesting an antibacterial mechanism that involves inhibition of cell division. Berberine is a DNA ligand and may induce filamentation through induction of the SOS response. Also, there is biochemical evidence for berberine inhibition of the cell division protein FtsZ. Here we aimed to assess possible berberine mechanism(s) of action in growing bacteria using genetics tools.Methodology/Principal FindingsFirst, we tested whether berberine inhibits bacterial growth through DNA damage and induction of the SOS response. The SOS response induced by berberine was much lower compared to that induced by mitomycin C in an SOS response reporter strain. Also, cell filamentation was observed in an SOS-negative E. coli strain. To test whether berberine inhibits FtsZ, we assessed its effects on formation of the cell division Z-rings, and observed a dramatic reduction in Z-rings in the presence of berberine. We next used two different strategies for RNA silencing of ftsZ and both resulted in sensitisation of bacteria to berberine, visible as a drop in the Minimum Inhibitory Concentration (MIC). Furthermore, Fractional Inhibitory Concentration Indices (FICIs) showed a high level of synergy between ftsZ silencing and berberine treatment (FICI values of 0.23 and 0.25 for peptide nucleic acid- and expressed antisense RNA-based silencing of ftsZ, respectively). Finally, over-expression of ftsZ led to a mild rescue effect in berberine-treated cells.ConclusionsThe results argue against DNA binding as the primary mechanism of action of berberine and support the hypothesis that its antibacterial properties are due to inhibition of the cell division protein FtsZ. In addition, the genetic approach used here provides a means to rapidly test the activity of other putative FtsZ inhibitors.

Highlights

Berberine (Figure 1) is an alkaloid produced by several plant species including barberry (Berberis spp.) and goldenseal (Hydrastis spp.), and is usually found in roots, stems and bark
The results argue against DNA binding as the primary mechanism of action of berberine and support the hypothesis that its antibacterial properties are due to inhibition of the cell division protein FtsZ
Berberine has been reported to be a DNA ligand and some of the effects it has on bacterial cells, cell elongation could be explained by DNA binding/damage and subsequent induction of the bacterial SOS response

Summary

Introduction

Berberine (Figure 1) is an alkaloid produced by several plant species including barberry (Berberis spp.) and goldenseal (Hydrastis spp.), and is usually found in roots, stems and bark. Berberine has been shown to exhibit antibacterial activity against a variety of bacteria, including many pathogenic species and multidrugresistant (MDR) strains of M. tuberculosis and MRSA. It is active against some fungi and protozoans. Berberine is a relatively weak antimicrobial, especially against Gram-negative bacteria. This is due to it being a substrate for the multi-drug resistance efflux pumps; the presence of MDR pump inhibitors remarkably increases the antibacterial effectiveness of the compound [10]. Escherichia coli exposed to berberine form filaments, suggesting an antibacterial mechanism that involves inhibition of cell division. We aimed to assess possible berberine mechanism(s) of action in growing bacteria using genetics tools

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Conclusion