Functional characterization of human osteoclast inhibitory peptide-1 (OIP-1/hSca) gene promoter.
We have recently identified and characterized the human osteoclast (OCL) inhibitory peptide-1 (OIP-1/hSca), a member of Ly-6 gene family. OIP-1 is an important physiologic regulator of OCL development and bone resorption activity. To determine the molecular mechanisms that regulate OIP-1 gene expression in OCL precursor cells, we isolated and characterized the OIP-1/hSca gene (2 Kb) promoter sequence. IFN-gamma (50 ng/ml) treatment of RAW 264.7 macrophage cells transfected with OIP-1 gene (-1 to -1988 bp relative to transcription start site) promoter-luciferase reporter plasmid demonstrated a significant (4 fold) increase in OIP-1 gene promoter activity. Sequence analysis of OIP-1 gene promoter region further identified a potential Stat-1 binding motif at -1629 to -1639 bp position. Stat-1 specific inhibitor, fludarabine (50 muM) abolished IFN-gamma stimulated OIP-1 gene promoter activity. Electrophoretic mobility shift assay (EMSA) further confirmed activated Stat-1 binding to the OIP-1 gene promoter sequence suggesting that IFN-gamma regulates OIP-1 gene expression in OCL precursor cells through a Stat-1 dependent signaling pathway. We further show that knock-down of TRADD enhances IFN-gamma signaling to increase OIP-1 gene expression in OCL precursor cells. These results should provide insights into the molecular control of OIP-1 gene expression and inhibition of OCL activity in the bone microenvironment.
Srinivasan
Shanmugarajan
S
Children's Research Institute, Medical University of South Carolina, 173 Ashley Avenue, Charleston, SC 29425, USA.
Ito
Masahiro
M
Kajiya
Hiroshi
H
Key
L Lyndon
LL
Johnson-Pais
Theresa L
TL
Reddy
Sakamuri V
SV
D000074183:ATPases Associated with Diverse Cellular Activities; D048868:Adaptor Proteins, Signal Transducing / Q000096:biosynthesis* / Q000235:genetics; D000818:Animals; D000970:Antineoplastic Agents / Q000378:metabolism / Q000494:pharmacology; D001853:Bone Marrow / Q000378:metabolism; D001862:Bone Resorption / Q000235:genetics / Q000378:metabolism*; D002454:Cell Differentiation / Q000502:physiology*; D002460:Cell Line; D024202:Electrophoretic Mobility Shift Assay / Q000379:methods; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology*; D007371:Interferon-gamma / Q000378:metabolism / Q000494:pharmacology; D060588:LIM Domain Proteins; D051379:Mice; D010010:Osteoclasts / Q000166:cytology / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex; D020218:Response Elements / Q000502:physiology*; D050794:STAT1 Transcription Factor / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D014157:Transcription Factors / Q000096:biosynthesis* / Q000235:genetics; D014740:Vidarabine / Q000031:analogs & derivatives / Q000494:pharmacology
D048868:Adaptor Proteins, Signal Transducing; D000970:Antineoplastic Agents; D060588:LIM Domain Proteins; C092282:PSMC5 protein, human; D050794:STAT1 Transcription Factor; C494079:STAT1 protein, human; D014157:Transcription Factors; D007371:Interferon-gamma; D046988:Proteasome Endopeptidase Complex; D000074183:ATPases Associated with Diverse Cellular Activities; D014740:Vidarabine; C024352:fludarabine