LEGIONELLA-PNEUMOPHILA HEAT-SHOCK PROTEIN-INDUCED INCREASE OF INTERLEUKIN-1-BETA MESSENGER-RNA INVOLVES PROTEIN-KINASE-C SIGNALING IN MACROPHAGES

Heat-shock proteins (hsp) are chaperon molecules important in protein Folding and assembly. Furthermore, they may have functions in immunore gulatory 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 macro phage immunity by directly increasing cytokine production. In the pres ent study we showed that the 60000 MW hsp of Legionella pneumophila (L p-hsp 60) increased cellular steady-state levels of interleukin-1 beta (IL-1 beta) mRNA measured by quantitative reverse transcription-polym erase chain reaction and Northern blotting as well as IL-1 secretion, when added to cultures of thioglycollate-elicited mouse peritoneal mac rophages in vitro. The level of mRNA increased in a dose-dependent man ner with a minimum effective concentration of 0.5 mu g/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 b ead uptake but permits binding, indicating that binding to the macroph age surface was sufficient for induction. Accumulation of IL-1 beta mR NA was completely blocked by pretreatment with the protein kinase C (P KC) inhibitor, H7, but not decreased by prior treatment with cyclohexi mide. The cell lysates of macrophages stimulated with Lp-hsp 60 showed an increased PKC activity measured by phosphorylation of PKC pseudosu bstrate. The IL-1 bioactivity in culture supernatants after 24 hr of s timulation with Lp-hsp 60 was increased in a dose-dependent manner but at hsp concentrations in excess of those needed to increase mRNA. Thu s, 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.