A Hemidominant Naip5 Allele in Mouse Strain MOLF/Ei-Derived Macrophages Restricts Legionella pneumophila Intracellular Growth

Abstract

Mouse-derived macrophages have the unique ability to restrict or permit Legionella pneumophila intracellular growth. The common inbred mouse strain C57BL/6J (B6) restricts L. pneumophila growth, whereas macrophages derived from A/J mice allow >10 3 -fold bacterial growth within three days. This phenotypic difference was mapped to the mouse Naip5 allele. The B6 restrictive Naip5 allele is dominant, and six amino acid changes in its product were predicted to control permissiveness. By using the wild-derived mouse strain MOLF/Ei, we found that MOLF/Ei-derived macrophages also restrict L. pneumophila growth, yet the Naip5 protein is identical to the A/J Naip5 at the six-amino-acid signature. The MOLF/Ei restrictive trait, unlike that of B6-derived macrophages, was not dominant over the A/J trait. In spite of this phenotypic difference, the L. pneumophila growth restriction in MOLF/Ei macrophages was mapped to the Naip5 region as well, indicating that the originally predicted change in the A/J Naip5 allele may not be critical for restriction. In the product of the A/J Naip5 permissive allele, there are four unique amino acid changes that map to a NACHT-like domain. Similar misregulating mutations have been identified in the NACHT domains of Nod-like receptor (NLR) proteins. Therefore, one of these mutations may be critical for restriction of L. pneumophila intracellular growth, and this parallels results found with human NLR variants with defects in the innate immune response.

The human apoptosis inhibitor NAIP induces pyroptosis in macrophages infected with Legionella pneumophila

“…Indeed, we could not observe L. pneumophila FlaAdependent responses in U937-derived macrophages (data not shown), which is in agreement with the previous report [23]. Considering that differences in the expression levels of mNaip5 among mouse strains affect the restriction of L. pneumophila growth in mBMMs [10,36], restoration of the factors could affect the susceptibility of human cells to L. pneumophila infection, as in our results. Indeed, transient overexpression of either human NAIP or NLRC4 has been shown to restrict L. pneumophila growth in human A549 cells, although the underlying mechanism has not been elucidated [27].…”

“…These results support the permissiveness to L. pneumophila of mBMMs derived from BALB/c mice [10], from which RAW264.7 cells are generated. It has been suggested that the differences in amino acid sequences and expression levels of mNaip5 among mouse strains affect susceptibility to L. pneumophila in mBMMs [10,36]. Indeed, although the nucleotide sequence of endogenous mNaip5 cDNA was intact (data not shown), expression of endogenous mNaip5 could not be detected by Western blotting of cell lysates from unmodified RAW264.7 cells using commercially available antibodies, which could not distinguish mNaip5 from other mNaip proteins based on their size (1402 aa), except for mNaip2 (1444 aa) (data not shown).…”

Expression and protective role of two novel NACHT-containing proteins in pathogen infection

“…Previous studies have focused on the critical role played by NACHT-family in the innate defense against bacterial infection (Franchi et al, 2006;Delbridge and O'Riordan, 2007;Vinzing et al, 2008;Losick et al, 2009;Koizumi et al, 2012;Kofoed et al, 2012). The human orthologs hNAIP and hNLRC4 were shown to inhibit of intracellular Legionella replication (Vinzing et al, 2008).…”

IFN-γ induction by neutrophil-derived IL-17A homodimer augments pulmonary antibacterial defense

“…The intracellular pathogen Legionella pneumophila causes both nosocomial and community pneumonia termed Legionnaires’ disease 2, 3 . In addition to the mouse model of L. pneumophila 4 infection, which mimics Legionnaires ' disease in immunocompetent individuals, 5, 6 macrophages from the inbred A/J mouse strain are often used in studies as they are permissive for growth of L. pneumophila 6, 7 whereas macrophages from other inbred mice are not 4 .…”