Helioxanthin inhibits interleukin-1 beta-induced MIP-1 beta production by reduction of c-jun expression and binding of the c-jun/CREB1 complex to the AP-1/CRE site of the MIP-1 beta promoter in Huh7 cells.
An elevated level of macrophage inflammatory protein-1beta (MIP-1beta) induced by IL-1beta has been correlated with chronic hepatic inflammatory disease. However, molecular mechanism of IL-1beta-induced MIP-1beta expression in hepatic cells is obscure. Previously, we reported the mechanism of the anti-hepatitis B virus (HBV) activity of helioxanthin (HE-145). Here, we demonstrated that HE-145 inhibited IL-1beta-induced MIP-1beta expression in a dose-dependent manner in Huh7 cells. To understand the mode of action of HE-145, we first examined how IL-1beta induced MIP-1beta expression at the molecular level. Using selective inhibitors, we found that JNK and p38 pathways participated in IL-1beta-induced MIP-1beta expression. HE-145 specifically suppressed IL-1beta-induced c-jun mRNA and protein expression and prevented c-jun-mediated AP-1 DNA-binding activity, whereas it had no effect on IL-1beta-induced activation of JNK, p38 and ATF2. Further studies indicated that HE-145 may downregulate c-jun mRNA expression directly at transcriptional level without requirement of de novo protein synthesis. Mutational analysis and supershift assays indicated that IL-1beta stimulated c-jun and CREB1 binding to the essential AP-1/CRE site of the MIP-1beta promoter. The inhibitory effect of HE-145 on IL-1beta-induced MIP-1beta promoter activity was completely reversed by overexpressing c-jun. Electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) assay consistently revealed that HE-145 reduced c-jun binding to the AP-1/CRE site in vitro and in vivo. Our results established a major role for c-jun in IL-1beta-induced MIP-1beta expression in hepatic cells. The reduction in IL-1beta-induced c-jun expression and subsequent binding of the c-jun/CREB1 complex to AP-1/CRE site mainly contributed to the inhibitory action of HE-145 on IL-1beta-induced MIP-1beta production.
Tseng
Pei-Chi
PC
Institute of Biochemistry and Molecular Biology, National Yang-Ming University, Taipei City 112, Taiwan, ROC.
Hsu
Hsing-Chih
HC
Janmanchi
Damodar
D
Lin
Chih-Hsiu
CH
Kuo
Yueh-Hsiung
YH
Chou
Chen-Kung
CK
Yeh
Sheau-Farn
SF
D045744:Cell Line, Tumor; D054407:Chemokine CCL4 / Q000037:antagonists & inhibitors* / Q000096:biosynthesis* / Q000235:genetics; D017362:Cyclic AMP Response Element-Binding Protein / Q000037:antagonists & inhibitors* / Q000378:metabolism; D051699:Cyclic AMP Response Element-Binding Protein A / Q000235:genetics / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects / Q000502:physiology; D006801:Humans; D053583:Interleukin-1beta / Q000037:antagonists & inhibitors* / Q000502:physiology*; D048031:JNK Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D017705:Lignans / Q000494:pharmacology*; D011401:Promoter Regions, Genetic / Q000187:drug effects / Q000502:physiology; D011485:Protein Binding / Q000187:drug effects / Q000502:physiology; D018808:Transcription Factor AP-1 / Q000235:genetics / Q000378:metabolism*
C115163:CREB1 protein, human; D054407:Chemokine CCL4; D017362:Cyclic AMP Response Element-Binding Protein; D051699:Cyclic AMP Response Element-Binding Protein A; D053583:Interleukin-1beta; D017705:Lignans; D018808:Transcription Factor AP-1; C505802:helioxanthin; D048031:JNK Mitogen-Activated Protein Kinases