Intracellular protein phosphorylation in murine peritoneal macrophages in response to bacterial lipopolysaccharide (LPS): effects of kinase-inhibitors and LPS-induced tolerance.
Intracellular protein phosphorylation is thought to be the initial step in cell activation. Bacterial lipopolysaccharide (LPS) induces a special set of the protein phosphorylation in the murine peritoneal macrophages, including p65 (molecular mass of 65 kDa) which is a substrate of serine kinase and the most dominant phosphorylated cytosolic protein. This article deals with the relation between the LPS-induced protein phosphorylation in the murine peritoneal macrophages and their productions of IL-1 beta and TNF-alpha. LPS-induced p65 phosphorylation seems to be dependent on protein kinase C (PKC) and calmodulin (CaM), because it diminishes in the presence of inhibitors to PKC or CaM. Tyrosine kinase inhibitors do not affect the p65 phosphorylation. The PKC inhibitors also affect the mRNA expressions and the productions of active molecules of IL-1 beta and TNF-alpha. Though the CaM inhibitor inhibits the mRNA expression and the active molecule production of IL-1 beta, it does not affect those of TNF-alpha. These results suggest that LPS-induced p65 phosphorylation is closely related to PKC and CaM, and that IL-1 beta production depends on PKC and CaM, while the TNF-alpha production is not dependent on CaM. These findings indicate the existence of multiple pathways and different regulatory mechanisms for transduction of LPS signal in the macrophages. Furthermore, LPS-induced phosphorylation is not observed in endotoxin tolerant macrophages after re-stimulation with LPS, suggesting that the LPS-stimulus signal is blocked at a site in the signal transduction-pathway before the point of phosphorylation of proteins in the tolerant macrophages.
Nakano
M
M
Department of Microbiology, Jichi Medical School, Tochigi, Japan.
Saito
S
S
Nakano
Y
Y
Yamasu
H
H
Matsuura
M
M
Shinomiya
H
H
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D003598:Cytoskeletal Proteins; D007546:Isoquinolines / Q000494:pharmacology; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D008264:Macrophages / Q000378:metabolism*; D051379:Mice; D008809:Mice, Inbred C3H; D008840:Microfilament Proteins; D010529:Peritoneal Cavity / Q000166:cytology; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D010879:Piperazines / Q000494:pharmacology; D047428:Protein Kinase Inhibitors; D011971:Receptors, Immunologic / Q000378:metabolism; D015398:Signal Transduction; D019311:Staurosporine
D000470:Alkaloids; D003598:Cytoskeletal Proteins; D007546:Isoquinolines; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D008840:Microfilament Proteins; D010750:Phosphoproteins; D010879:Piperazines; D047428:Protein Kinase Inhibitors; D011971:Receptors, Immunologic; C066559:Lcp1 protein, mouse; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D019311:Staurosporine