AUTHOR=Panta Pradip R. , Doerrler William T. 

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

JOURNAL=Frontiers in Microbiology

VOLUME=11

YEAR=2021

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2020.618389

DOI=10.3389/fmicb.2020.618389

ISSN=1664-302X

ABSTRACT=<p>The DedA family is a conserved membrane protein family found in most organisms. A <italic>Burkholderia thailandensis</italic> 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. <italic>B. thailandensis</italic> Δ<italic>dbcA</italic> lipid A contains only small amounts of Ara4N, likely leading to colistin sensitivity. Two <italic>B. thailandensis</italic> 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 <italic>arn</italic> operon by genomic insertion of inducible promoters. We found that overexpression of <italic>arn</italic> operons in Δ<italic>dbcA</italic> 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 Δ<italic>dbcA</italic>. In addition, the products of <italic>arn</italic> operons are essential for acid tolerance, suggesting a physiological function of Ara4N modification. Finally, we show that Δ<italic>dbcA</italic> is sensitive to bacitracin and expression of a <italic>B. thailandensis</italic> UppP/BacA homolog (<italic>BTH_I1512)</italic> can partially restore resistance to bacitracin. Expression of a different UppP/BacA homolog (<italic>BTH_I2750)</italic> 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 <italic>B. thailandensis.</italic></p>