NF-kappaB-dependent induction of cathelicidin-related antimicrobial peptide in murine mast cells by lipopolysaccharide.
BACKGROUND
An important aspect of the innate immune response to pathogens is the production of anti-microbial peptides such as cathelicidin-related antimicrobial peptide (CRAMP), the murine homologue of human cathelicidin LL-37. In this study, mechanisms regulating LPS-induction of CRAMP gene expression in mast cells were investigated. NF-kappaB and MAPK pathways were the focus of investigation.


METHODS
Mouse bone marrow-derived mast cells were grown in culture and stimulated with LPS. MAPKs and NF-kappaB were monitored by immunoblot analysis. ERK, JNK and p38 MAPK were inhibited using siRNAs or a pharmacological inhibitor. Accumulation of the p65 component of NF-kappaB was inhibited by siRNA and NF-kappaB activation was inhibited by overexpression of I kappaB alpha. MEKK2 or MEKK3 were overexpressed by transfection. The effects of all of these treatments on CRAMP gene expression were monitored by RT-PCR.


RESULTS
Inhibition of ERK, JNK or p38 MAPK had little discernible effect on LPS-inducible CRAMP gene expression. Overexpression of MEKK2 or MEKK3 likewise had little impact. However, inhibition of the accumulation of p65 NF-kappaB prevented LPS-induced CRAMP mRNA. An important role for NF-kappaB in CRAMP gene expression was confirmed by overexpression of I kappaB alpha, which reduced both basal and induced levels of CRAMP mRNA.


CONCLUSIONS
NF-kappaB, but not MAPKs, plays an important role in LPS-mediated induction of CRAMP gene in mast cells. Defects which inhibit NF-kappaB activity may increase susceptibility to bacterial and viral pathogens which are sensitive to cathelicidins.
Li
Guiming
G
Program in Cell Biology, Department of Pediatrics, National Jewish Health, Denver, CO 80206, USA.
Domenico
Joanne
J
Jia
Yi
Y
Lucas
Joseph J
JJ
Gelfand
Erwin W
EW
D000818:Animals; D023181:Antimicrobial Cationic Peptides / Q000235:genetics*; D001854:Bone Marrow Cells / Q000166:cytology; D054804:Cathelicidins; D004305:Dose-Response Relationship, Drug; D015870:Gene Expression / Q000187:drug effects / Q000235:genetics; D005786:Gene Expression Regulation / Q000502:physiology*; D020672:I-kappa B Proteins / Q000235:genetics / Q000378:metabolism; D048031:JNK Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D048730:MAP Kinase Kinase Kinase 2 / Q000235:genetics; D048748:MAP Kinase Kinase Kinase 3 / Q000235:genetics; D008407:Mast Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D034741:RNA, Small Interfering / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D051197:Toll-Like Receptor 4 / Q000378:metabolism; D051996:Transcription Factor RelA / Q000235:genetics; D014162:Transfection; D048051:p38 Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism
D023181:Antimicrobial Cationic Peptides; D054804:Cathelicidins; D020672:I-kappa B Proteins; D008070:Lipopolysaccharides; D016328:NF-kappa B; C000606204:NFKBIA protein, human; C000606205:Nfkbia protein, mouse; D034741:RNA, Small Interfering; C493487:Tlr4 protein, mouse; D051197:Toll-Like Receptor 4; D051996:Transcription Factor RelA; D000072000:NF-KappaB Inhibitor alpha; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D048730:MAP Kinase Kinase Kinase 2; D048748:MAP Kinase Kinase Kinase 3; C482619:Map3k2 protein, mouse; C482626:Map3k3 protein, mouse