Tosylphenylalanine chloromethyl ketone inhibits TNF-alpha mRNA synthesis in the presence of activated NF-kappa B in RAW 264.7 macrophages.
Serine proteinase inhibitors such as N-tosyl-L-phenylalanine chloromethyl ketone (TPCK) and N alpha-p-tosyl-L-lysine chloromethyl ketone (TLCK) were shown to inhibit production of tumour necrosis factor-alpha (TNF-alpha) in lipopolysaccharide (LPS)-activated RAW 264.7 macrophages. The proteinase inhibitors were also reported to inhibit activation of the transcription factor nuclear factor-kappa B (NF-kappa B) by blocking the signalling pathway for stimuli-induced phosphorylation of the inhibitory subunit (I kappa B alpha) and thus preventing its degradation. In RAW 264.7 cells TPCK and TLCK significantly suppressed LPS-induced increase in TNF-alpha mRNA, induction of nuclear kappa B-binding activity and degradation of I kappa B alpha. TPCK and TLCK effectively blocked TNF-alpha mRNA synthesis even when they were added after LPS stimulation. In these cells, however, the inhibitory modes of the two inhibitors were found to be different: while addition of TLCK suppressed I kappa B alpha degradation and reduced NF-kappa B activity, a comparable decrease in the nuclear kappa B-binding activity or in I kappa B alpha degradation was not observed in cells treated with TPCK. Our results show that TPCK inhibits LPS-induced TNF-alpha mRNA synthesis in the presence of activated NF-kappa B and suggests that mechanisms other than NF-kappa B activation are involved in the transcriptional regulation of the TNF-alpha gene.
Jeong
J Y
JY
Department of Biochemistry, Catholic University Medical College, Seoul, South Korea.
Kim
K U
KU
Jue
D M
DM
D000818:Animals; D015152:Blotting, Northern; D018929:Cell Culture Techniques; D008070:Lipopolysaccharides / Q000276:immunology; D008262:Macrophage Activation / Q000276:immunology*; D008264:Macrophages / Q000276:immunology*; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics; D014107:Tosyllysine Chloromethyl Ketone / Q000494:pharmacology; D014108:Tosylphenylalanyl Chloromethyl Ketone / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis* / Q000187:drug effects / Q000235:genetics
D008070:Lipopolysaccharides; D016328:NF-kappa B; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha; D014107:Tosyllysine Chloromethyl Ketone; D014108:Tosylphenylalanyl Chloromethyl Ketone