The Bacillus anthracis protein MprF is required for synthesis of lysylphosphatidylglycerols and for resistance to cationic antimicrobial peptides.
During inhalational anthrax, Bacillus anthracis survives and replicates in alveolar macrophages, followed by rapid invasion into the host's bloodstream, where it multiplies to cause heavy bacteremia. B. anthracis must therefore defend itself from host immune functions encountered during both the intracellular and the extracellular stages of anthrax infection. In both of these niches, cationic antimicrobial peptides are an essential component of the host's innate immune response that targets B. anthracis. However, the genetic determinants of B. anthracis contributing to resistance to these peptides are largely unknown. Here we generated Tn917 transposon mutants in the DeltaANR strain (pXO1- pXO2-) of B. anthracis and screened them for altered protamine susceptibility. A protamine-sensitive mutant identified carried the transposon inserted in the BA1486 gene encoding a putative membrane protein homologous to MprF known in several gram-positive pathogens. A mutant strain with the BAS1375 gene (the orthologue of BA1486) deleted in the Sterne 34F2 strain (pXO1+ pXO2-) of B. anthracis exhibited hypersusceptibility not only to protamine but also to alpha-helical cathelicidin LL-37 and beta-sheet defensin human neutrophil peptide 1 compared to the wild-type Sterne strain. Analysis of membrane lipids using isotopic labeling demonstrated that the BAS1375 deletion mutant is unable to synthesize lysinylated phosphatidylglycerols, and this defect is rescued by genetic complementation. Further, we determined the structures of these lysylphosphatidylglycerols by using various mass spectrometric analyses. These results demonstrate that in B. anthracis a functional MprF is required for the biosynthesis of lysylphosphatidylglycerols, which is critical for resistance to cationic antimicrobial peptides.
Samant
Shalaka
S
Center for Pharmaceutical Biotechnology, University of Illinois at Chicago, M/C 870, 900 S. Ashland Ave., Chicago, Illinois 60607, USA.
Hsu
Fong-Fu
FF
Neyfakh
Alexander A
AA
Lee
Hyunwoo
H
D023181:Antimicrobial Cationic Peptides / Q000494:pharmacology*; D001408:Bacillus anthracis / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D024881:Drug Resistance, Bacterial; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D005798:Genes, Bacterial; D008239:Lysine / Q000096:biosynthesis*; D010715:Phosphatidylglycerols / Q000096:biosynthesis*; D011479:Protamines / Q000494:pharmacology
D023181:Antimicrobial Cationic Peptides; D001426:Bacterial Proteins; D010715:Phosphatidylglycerols; D011479:Protamines; C002285:lysylphosphatidylglycerol; D008239:Lysine