Inhibition of lipopolysaccharide-induced macrophage IL-12 production by Leishmania mexicana amastigotes: the role of cysteine peptidases and the NF-kappaB signaling pathway.
Infection with lesion-derived Leishmania mexicana amastigotes inhibited LPS-induced IL-12 production by mouse bone marrow-derived macrophages. This effect was associated with expression of cysteine peptidase B (CPB) because amastigotes of CPB deletion mutants had limited ability to inhibit IL-12 production, whereas preincubation of cells with a CPB inhibitor, cathepsin inhibitor IV, was able to suppress the effect of wild-type amastigotes. Infection with wild-type amastigotes resulted in a time-dependent proteolytic degradation of IkappaBalpha and IkappaBbeta and the related protein NF-kappaB. This effect did not occur with amastigotes of CPB deletion mutants or wild-type promastigotes, which do not express detectable CPB. NF-kappaB DNA binding was also inhibited by amastigote infection, although nuclear translocation of cleaved fragments of p65 NF-kappaB was still observed. Cysteine peptidase inhibitors prevented IkappaBalpha, IkappaBbeta, and NF-kappaB degradation induced by amastigotes, and recombinant CPB2.8, an amastigote-specific isoenzyme of CPB, was shown to degrade GST-IkappaBalpha in vitro. LPS-mediated IkappaBalpha and IkappaBbeta degradation was not affected by these inhibitors, confirming that the site of degradation of IkappaBalpha, IkappaBbeta, and NF-kappaB by the amastigotes was not receptor-driven, proteosomal-mediated cleavage. Infection of bone marrow macrophages with amastigotes resulted in cleavage of JNK and ERK, but not p38 MAPK, whereas preincubation with a cysteine peptidase inhibitor prevented degradation of these proteins, but did not result in enhanced protein kinase activation. Collectively, our results suggest that the amastigote-specific cysteine peptidases of L. mexicana are central to the ability of the parasite to modulate signaling via NF-kappaB and consequently inhibit IL-12 production.
Cameron
Pamela
P
Department of Immunology, Strathclyde Institute for Biomedical Sciences, University of Strathclyde, Glasgow, United Kingdom.
McGachy
Adrienne
A
Anderson
Mary
M
Paul
Andrew
A
Coombs
Graham H
GH
Mottram
Jeremy C
JC
Alexander
James
J
Plevin
Robin
R
D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000276:immunology / Q000378:metabolism*; D004247:DNA / Q000378:metabolism; D020672:I-kappa B Proteins / Q000378:metabolism; D018664:Interleukin-12 / Q000096:biosynthesis* / Q000276:immunology; D007894:Leishmania mexicana / Q000276:immunology / Q000378:metabolism; D007896:Leishmaniasis / Q000276:immunology* / Q000378:metabolism; D008070:Lipopolysaccharides / Q000276:immunology*; D008264:Macrophages / Q000276:immunology / Q000378:metabolism*; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D016328:NF-kappa B / Q000276:immunology / Q000378:metabolism*
D020672:I-kappa B Proteins; D008070:Lipopolysaccharides; D016328:NF-kappa B; D018664:Interleukin-12; D004247:DNA; D020928:Mitogen-Activated Protein Kinases; D003546:Cysteine Endopeptidases