Defective translation of tumor necrosis factor mRNA in lipopolysaccharide-tolerant macrophages.
Macrophage activation by lipopolysaccharide (LPS) results in the translational activation of tumor necrosis factor (TNF) mRNA. The initial phase of macrophage activation is followed by a refractory state called LPS tolerance characterized by an impaired TNF production in response to a secondary LPS challenge. LPS-tolerant macrophages contain high amounts of TNF mRNA, suggesting a translational regulation of TNF biosynthesis. The induction of LPS tolerance was studied in RAW 264.7 macrophages stably transfected with a chloramphenicol acetyl-transferase (CAT) reporter gene construct driven by a constitutive cytomegalovirus promoter and containing the 3' untranslated region of the murine TNF gene. We found that primary stimulation of transfected cells by LPS (1 ng/ml, 12 hr) resulted in a marked suppression (80%) of CAT accumulation in response to a secondary LPS challenge (1 microgram/ml, 6 hr). In contrast, the accumulation of CAT mRNA was not influenced by LPS tolerance. Using the same CAT reporter, we observed that the serine/threonine phosphatases 1 and 2A inhibitor okadaic acid induced TNF mRNA translation and that this activation was not inhibited by LPS-tolerance. In conclusion, these data indicate that deficient production of TNF in LPS-tolerant macrophages in response to a second LPS challenge is characterized by a defective translation of TNF mRNA. However, this hyporesponsiveness to LPS is specific, since translation of TNF mRNA induced by okadaic acid is not inhibited in LPS-tolerant macrophages.
Marchant
A
A
Service d'Immunologie, Hôpital Erasme, Université Libre de Bruxelles, Belgium.
Gueydan
C
C
Houzet
L
L
Amraoui
Z
Z
Sels
A
A
Huez
G
G
Goldman
M
M
Kruys
V
V
D000818:Animals; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D003587:Cytomegalovirus / Q000235:genetics; D004361:Drug Tolerance; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004988:Ethers, Cyclic / Q000494:pharmacology; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D007700:Kinetics; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008262:Macrophage Activation; D008264:Macrophages / Q000378:metabolism*; D051379:Mice; D019319:Okadaic Acid; D010744:Phosphoric Monoester Hydrolases / Q000037:antagonists & inhibitors; D011401:Promoter Regions, Genetic; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics* / Q000378:metabolism; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis / Q000235:genetics*
D004791:Enzyme Inhibitors; D004988:Ethers, Cyclic; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D019319:Okadaic Acid; D015500:Chloramphenicol O-Acetyltransferase; D010744:Phosphoric Monoester Hydrolases