# A lignan compound regulates LPS modifications via PmrA/B signaling cascades to potentiate colistin efficacy in vivo

## TL;DR

A natural lignan compound called magnolol enhances the effectiveness of the antibiotic colistin by blocking bacterial defenses, offering a new way to combat antibiotic resistance.

## Contribution

The discovery of magnolol as a novel adjuvant that potentiates colistin efficacy by inactivating PmrA/B signaling in Gram-negative bacteria.

## Key findings

- Magnolol disrupts PmrA/B signaling by dissociating PmrA from its DNA targets, blocking LPS modifications.
- This action increases colistin binding to bacterial membranes, leading to rapid cell death via pore formation.
- Phytochemicals like magnolol represent a new class of adjuvants to extend the lifespan of last-line antibiotics.

## Abstract

There has been a substantial gap between drying antibiotic pipeline and ongoing antibiotic resistance crisis, necessitating approaches to revitalize existing antimicrobials to meet unmet clinical demand for viable treatments. Herein, a lignan compound, magnolol, was identified that profoundly potentiates colistin (CS) to eradicate Gram negative bacteria and curb the development of resistance under host-mimicking condition. The mechanistic study showed that magnolol is able to disrupt PmrA/B two component signaling by dissociating the PmrA regulator protein from its cognate DNA including eptA and arnT. This action blocks the PmrA/B-dependent protective modifications of lipopolysaccharide (LPS) to reduce the net charges of bacterial membrane, thereaby facilitating its electrostatic interaction with CS. MAG-facilitated enhancement of CS binding promotes the formation of toroidal pores in the bacterial membrane, which in turn triggers rapid bacterial death by inducing lethal cytoplasmic contents leakage. In sum, this work not only illustrates the great potential of untapped phytoconstitutes such as magnolol in confronting antibiotic resistance but also reveals that silencing PmrA/B signaling as a favorable strategy to potentiate CS activity in vivo.

As one of the last-line antibiotics, the colistin holds clinically indispensable for treating against MDR Gram negative pathogens but is limited by toxicity and growing resistance. Importantly, certain pathogens can develop colistin resistance in response to in-host stimuli, thereby offsetting colistin efficacy in vivo. In this context, well-designed adjuvants that potentiate its activity in vivo allow colistin to be used at lower, safer and more precise doses, and help extend the lifespan of this last-line therapy. Herein, we applied a high throughput screening based on a host-mimicking LPM medium and identified a lignan compound, magnolol (MAG), that is able to potentiate colistin efficacy in vitro and in vivo. It was found that this potentiation relies on MAG’s ability to paralyze PmrA/B-dependent membrane modifications to facilitate the colistin binding and destabilizing bacterial membrane. We also propose a new mode of PmrA/B inactivation for MAG, in which it acts as a molecular decoy to dissociate the PmrA regulator protein from its target DNA. These findings suggest that the phytochemicals, exemplified by the MAG, are untapped source for potential antibiotic adjuvants and trapping the transcriptional regulation of the PmrA/B two component system may present a new mechanistic paradigm for developing next-generation colistin adjuvants.