A Burkholderia thailandensis DedA Family Membrane Protein Is Required for Proton Motive Force Dependent Lipid A Modification.

Pradip R Panta,William T Doerrler

doi:10.3389/fmicb.2020.618389

Abstract

The DedA family is a conserved membrane protein family found in most organisms. A Burkholderia thailandensis DedA family protein, named DbcA, is required for high-level colistin (polymyxin E) resistance, but the mechanism awaits elucidation. Modification of lipopolysaccharide lipid A with the cationic sugar aminoarabinose (Ara4N) is required for colistin resistance and is dependent upon protonmotive force (PMF) dependent transporters. B. thailandensis ΔdbcA lipid A contains only small amounts of Ara4N, likely leading to colistin sensitivity. Two B. thailandensis operons are required for lipid A modification with Ara4N, one needed for biosynthesis of undecaprenyl-P-Ara4N and one for transport of the lipid linked sugar and subsequent lipid A modification. Here, we directed overexpression of each arn operon by genomic insertion of inducible promoters. We found that overexpression of arn operons in ΔdbcA can partially, but not completely, restore Ara4N modification of lipid A and colistin resistance. Artificially increasing the PMF by lowering the pH of the growth media also increased membrane potential, amounts of Ara4N, and colistin resistance of ΔdbcA. In addition, the products of arn operons are essential for acid tolerance, suggesting a physiological function of Ara4N modification. Finally, we show that ΔdbcA is sensitive to bacitracin and expression of a B. thailandensis UppP/BacA homolog (BTH_I1512) can partially restore resistance to bacitracin. Expression of a different UppP/BacA homolog (BTH_I2750) can partially restore colistin resistance, without changing the lipid A profile. This work suggests that maintaining optimal membrane potential at slightly alkaline pH media by DbcA is responsible for proper modification of lipid A by Ara4N and provides evidence of lipid A modification-dependent and -independent mechanisms of colistin resistance in B. thailandensis.

Highlights

Colistin is a polycationic and amphiphilic peptide which is a last resort antibiotic for treatment of many Gram-negative bacterial infections
The increasing threat of multidrug-resistant (MDR) bacterial pathogens with stagnant discovery of new class of effective antibiotics has led to renewed interest in reviving older antibiotics like colistin, a polycationic cyclic polypeptide belonging to polymyxin family of antibiotics (Talbot et al, 2006)
Colistin has been classified as “Reserve Group” of antibiotics to be used as a lastresort antibiotic against MDR infections that cannot be treated by any other conventional antibiotics (Hsia et al, 2019)

Summary

Introduction

Colistin (polymyxin E) is a polycationic and amphiphilic peptide which is a last resort antibiotic for treatment of many Gram-negative bacterial infections. Clinical use of colistin is increasing due to limited alternative therapeutic options (Li et al, 2006). The emergence of colistin resistance has been documented in several human pathogens, such as K. pneumoniae (Bratu et al, 2005; Antoniadou et al, 2007; Farzana et al, 2020), Salmonella enterica serovar Newport (Elbediwi et al, 2020), and Acinetobacter baumannii (Marano et al, 2020; Papathanakos et al, 2020). More worrisome is the recent global spread of mobilized colistin resistance (mcr) genes (Elbediwi et al, 2019). Understanding the molecular mechanism of colistin resistance is of utmost importance at times when the antibiotic discovery pipeline is drying up

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