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The Ku Antigen-Recombination Signal-binding Protein Jκ Complex Binds to the Nuclear Factor-κB p50 Promoter and Acts as a Positive Regulator of p50 Expression in Human Gastric Cancer Cells

 Joo Weon Lim  ;  Hyeyoung Kim  ;  Kyung Hwan Kim 
 JOURNAL OF BIOLOGICAL CHEMISTRY, Vol.279(1) : 231-237, 2004 
Journal Title
Issue Date
Antigens, Nuclear/chemistry ; Antigens, Nuclear/genetics* ; Base Sequence ; Binding Sites ; DNA Helicases* ; DNA Primers ; DNA-Binding Proteins/chemistry ; DNA-Binding Proteins/genetics* ; DNA-Binding Proteins/metabolism* ; Humans ; Immunoglobulin J Recombination Signal Sequence-Binding Protein ; Ku Autoantigen ; NF-kappa B/genetics* ; Nuclear Proteins/metabolism* ; Oligonucleotide Probes ; Promoter Regions, Genetic/physiology* ; Protein Subunits/genetics ; Recombinases/metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Sequence Alignment ; Sequence Homology, Nucleic Acid ; Stomach Neoplasms ; Transcription Factors/genetics ; Transfection ; Tumor Cells, Cultured
The p50 subunit of NF-kappaB is a transcription factor that regulates the expression of a variety of genes. Previously, we showed that the expression of Ku antigen, a DNA repair protein, is mediated by NF-kappaB in gastric cancer AGS cells (Lim, J. W., Kim, H., and Kim, K. H. (2002) J. Biol. Chem. 277, 46093-46100). In this study, we report that the inhibition of Ku activity reduced both p50 expression and nuclear NF-kappaB activity in AGS cells. A co-immunoprecipitation experiment demonstrated that Ku antigen interacted with recombination signal-binding protein Jkappa (RBP-Jkappa), a DNA-binding protein. Ku antigen, RBP-Jkappa, and p50 were found to bind to the DNA region containing the kappaB element in the p50 promoter. Supershift and competition experiments demonstrated that Ku antigen and RBP-Jkappa bound sequence-specifically to downstream elements of kappaB at GCTTC and TGGGGG. mRNA expression and de novo synthesis of p50 were inhibited in cells transfected with the mutant gene expression constructs for IkappaBalpha, Ku80, and RBP-Jkappa. A reporter assay demonstrated that p50 transcription was positively mediated by NF-kappaB, Ku antigen, and RBP-Jkappa and that the binding elements for these proteins were required for optimal p50 expression. The interaction of Ku antigen with RBP-Jkappa and NF-kappaB p50 may act as a positive regulator of p50 expression in gastric cancer AGS cells.
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1. College of Medicine (의과대학) > Dept. of Pharmacology (약리학교실) > 1. Journal Papers
Yonsei Authors
Kim, Kyung Hwan(김경환)
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