RRC ID 67955
Author Ishii J, Kitazawa R, Mori K, McHugh KP, Morii E, Kondo T, Kitazawa S.
Title Lipopolysaccharide suppresses RANK gene expression in macrophages by down-regulating PU.1 and MITF.
Journal J Cell Biochem
Abstract Receptor activator of NF-kappaB (RANK) is a receptor for RANK ligand (RANKL), and signals transduced by RANK-RANKL interaction are prerequisite for the differentiation and activation of osteoclasts. We cloned and characterized a 6-kb fragment containing the 5'-flanking region of the mouse RANK gene. A fragment of 1-kb from the transcription start sites containing four Sp-1 sites and putative binding sites for MITF, CRE/AP-1, and PU.1 was ligated to the pGL3-basic vector, and the promoter activity was confirmed by transfection studies. By electrophoretic gel motility shift assay, both PU.1 and proximal MITF binding site showed specific DNA-protein binding. Co-transfection studies with MITF- and PU.1-expression vectors revealed that MITF and PU.1 increased RANK promoter activity three- and twofold, respectively, and sixfold synergistically. Taken together, these results show that RANK transcription is positively regulated by both PU.1 and MITF. The effect of lipopolysaccharide (LPS) on RANK gene expression, analyzed by in situ hybridization using mouse bone tissue, showed that LPS decreased RANK transcripts of both precursor and mature osteoclasts. Furthermore, LPS treatment of RAW.264.7 cells decreased their RANK mRNA expression by 70%, mirroring the decrease of PU.1 and MITF mRNA. Short-term treatment with LPS decreased the promoter activity of pGL3-WT by 70%. Although LPS has been reported to promote osteoclastogenesis in chronic and local pyogenic inflammation, we speculate that LPS per se may directly suppress RANK expression in the osteoclastic cell lineage by down-regulating the expression of PU.1 and MITF genes in acute and systemic severe endotoxemia, such as in septic shock.
Volume 105(3)
Pages 896-904
Published 2008-10-15
DOI 10.1002/jcb.21886
PMID 18759249
MeSH Animals Down-Regulation* Female Lipopolysaccharides / pharmacology* Macrophages / metabolism* Mice Mice, Inbred BALB C Microphthalmia-Associated Transcription Factor / genetics* Microphthalmia-Associated Transcription Factor / metabolism Osteoclasts / metabolism Promoter Regions, Genetic Proto-Oncogene Proteins / genetics* Proto-Oncogene Proteins / metabolism RNA, Messenger / metabolism Receptor Activator of Nuclear Factor-kappa B / genetics* Receptor Activator of Nuclear Factor-kappa B / metabolism Trans-Activators / genetics* Trans-Activators / metabolism Transfection
IF 4.237
Human and Animal Cells RAW 264(RCB0535)