title: Listeria monocytogenes is resistant to lysozyme through the regulation, not the acquisition, of cell wall-modifying enzymes.
abstract: Listeria monocytogenes is a Gram-positive facultative intracellular pathogen that is highly resistant to lysozyme, a ubiquitous enzyme of the innate immune system that degrades cell wall peptidoglycan. Two peptidoglycan-modifying enzymes, PgdA and OatA, confer lysozyme resistance on L. monocytogenes; however, these enzymes are also conserved among lysozyme-sensitive nonpathogens. We sought to identify additional factors responsible for lysozyme resistance in L. monocytogenes. A forward genetic screen for lysozyme-sensitive mutants led to the identification of 174 transposon insertion mutations that mapped to 13 individual genes. Four mutants were killed exclusively by lysozyme and not other cell wall-targeting molecules, including the peptidoglycan deacetylase encoded by pgdA, the putative carboxypeptidase encoded by pbpX, the orphan response regulator encoded by degU, and the highly abundant noncoding RNA encoded by rli31. Both degU and rli31 mutants had reduced expression of pbpX and pgdA, yet DegU and Rli31 did not regulate each other. Since pbpX and pgdA are also present in lysozyme-sensitive bacteria, this suggested that the acquisition of novel enzymes was not responsible for lysozyme resistance, but rather, the regulation of conserved enzymes by DegU and Rli31 conferred high lysozyme resistance. Each lysozyme-sensitive mutant exhibited attenuated virulence in mice, and a time course of infection revealed that the most lysozyme-sensitive strain was killed within 30 min of intravenous infection, a phenotype that was recapitulated in purified blood. Collectively, these data indicate that the genes required for lysozyme resistance are highly upregulated determinants of L. monocytogenes pathogenesis that are required for avoiding the enzymatic activity of lysozyme in the blood.
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, California, USA.
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Burke
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, California, USA.
authors.forename: Anastasia
authors.initials: A
authors.lastname: Loukitcheva
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, California, USA.
authors.forename: Jason
authors.initials: J
authors.lastname: Zemansky
authors.affiliation: Institut Pasteur, Biology and Genetics of the Bacterial Cell Wall Unit, Paris, France INSERM, Aveny Group, Paris, France.
authors.forename: Richard
authors.initials: R
authors.lastname: Wheeler
authors.affiliation: Institut Pasteur, Biology and Genetics of the Bacterial Cell Wall Unit, Paris, France INSERM, Aveny Group, Paris, France.
authors.forename: Ivo G
authors.initials: IG
authors.lastname: Boneca
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, California, USA School of Public Health, University of California, Berkeley, Berkeley, California, USA portnoy@berkeley.edu.
authors.forename: Daniel A
authors.initials: DA
authors.lastname: Portnoy
mesh_terms: D000581:Amidohydrolases / Q000378:metabolism; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D002268:Carboxypeptidases / Q000378:metabolism; D002473:Cell Wall / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D015971:Gene Expression Regulation, Enzymologic / Q000502:physiology*; D008089:Listeria monocytogenes / Q000166:cytology / Q000201:enzymology* / Q000378:metabolism / Q000472:pathogenicity; D008088:Listeriosis / Q000382:microbiology; D051379:Mice; D008810:Mice, Inbred C57BL; D008813:Mice, Inbred ICR; D009113:Muramidase / Q000378:metabolism*; D016254:Mutagenesis, Insertional; D010457:Peptidoglycan / Q000378:metabolism; D014774:Virulence; D047090:beta-Lactams / Q000494:pharmacology
chemical_list: D000900:Anti-Bacterial Agents; D010457:Peptidoglycan; D047090:beta-Lactams; D009113:Muramidase; D002268:Carboxypeptidases; D000581:Amidohydrolases