Legionella pneumophila heat-shock protein-induced increase of interleukin-1 beta mRNA involves protein kinase C signalling in macrophages.
Heat-shock proteins (hsp) are chaperon molecules important in protein folding and assembly. Furthermore, they may have functions in immunoregulatory processes, like T-cell stimulation and antigen presentation, which are not yet fully understood. It has been shown that several hsp of various species and family derivations modulate functions in macrophage immunity by directly increasing cytokine production. In the present study we showed that the 60,000 MW hsp of Legionella pneumophila (Lp-hsp 60) increased cellular steady-state levels of interleukin-1 beta (IL-1 beta) mRNA measured by quantitative reverse transcription-polymerase chain reaction and Northern blotting as well as IL-1 secretion, when added to cultures of thioglycollate-elicited mouse peritoneal macrophages in vitro. The level of mRNA increased in a dose-dependent manner with a minimum effective concentration of 0.5 microgram/ml and peaked 3 hr after stimulation. Lp-hsp 60-coated latex beads also increased IL-1 beta mRNA levels in the presence of cytochalasin D, which inhibits bead uptake but permits binding, indicating that binding to the macrophage surface was sufficient for induction. Accumulation of IL-1 beta mRNA was completely blocked by pretreatment with the protein kinase C (PKC) inhibitor, H7, but not decreased by prior treatment with cycloheximide. The cell lysates of macrophages stimulated with Lp-hsp 60 showed an increased PKC activity measured by phosphorylation of PKC pseudosubstrate. The IL-1 bioactivity in culture supernatants after 24 hr of stimulation with Lp-hsp 60 was increased in a dose-dependent manner but at hsp concentrations in excess of those needed to increase mRNA. Thus, the present study demonstrates that Lp-hsp 60 rapidly increases the steady-state level of IL-1 beta mRNA, possibly through a cell surface receptor system involving a PKC-dependent signalling pathway.
Retzlaff
C
C
Department of Medical Microbiology and Immunology, University of South Florida College of Medicine, Tampa 4799, USA.
Yamamoto
Y
Y
Okubo
S
S
Hoffman
P S
PS
Friedman
H
H
Klein
T W
TW
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000494:pharmacology; D000818:Animals; D001426:Bacterial Proteins / Q000494:pharmacology*; D015152:Blotting, Northern; D002478:Cells, Cultured; D018834:Chaperonin 60 / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D005260:Female; D007375:Interleukin-1 / Q000235:genetics / Q000378:metabolism*; D016952:Legionella pneumophila; D008264:Macrophages / Q000187:drug effects / Q000201:enzymology / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D016133:Polymerase Chain Reaction; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D015398:Signal Transduction / Q000502:physiology
D001426:Bacterial Proteins; D018834:Chaperonin 60; D007375:Interleukin-1; D012333:RNA, Messenger; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D011493:Protein Kinase C