Positive and negative regulation of nuclear factor-kappaB-mediated transcription by IkappaB-zeta, an inducible nuclear protein.
IkappaB-zeta is an inducible nuclear protein that interacts with nuclear factor-kappaB (NF-kappaB) via its carboxyl-terminal ankyrin-repeats. Previous studies using an NF-kappaB reporter have shown that IkappaB-zeta inhibits the activity of NF-kappaB. In the present study, we dissected the amino-terminal region of IkappaB-zeta, which shows no homology to any other proteins. Indirect immunofluorescence studies demonstrated the presence of a bipartite nuclear localization signal spanning amino acids 163-178. Using GAL4 fusion proteins, we found that internal fragments containing amino acids 329-402 possessed intrinsic transcriptional activation activity. Interestingly, the activity was not detected in GAL4 fusion proteins of the full-length IkappaB-zeta. On the other hand, the GAL4-dependent transcriptional activity was generated by co-expression of the GAL4-NF-kappaB p50 subunit fusion protein and the full-length IkappaB-zeta, neither of which exhibited the activity on their own. A new splicing variant, IkappaB-zeta(D), with a deletion of amino acids 236-429, was found to lack transactivation activity. Forced expression of IkappaB-zeta, but not IkappaB-zeta(D), augmented interleukin-6 production, indicating the functional significance of the transactivation activity. In contrast, tumor necrosis factor-alpha production was inhibited by expression of IkappaB-zeta, highlighting the dual functions of this molecule. These results indicate that IkappaB-zeta harbors latent transcriptional activation activity, and that the activity is expressed upon interaction with the NF-kappaB p50 subunit. In addition to the inhibitory activity on NF-kappaB-mediated transcription, the transcriptional activation activity of IkappaB-zeta should be crucial for the regulation of inflammation.
Motoyama
Masaiwa
M
Department of Molecular and Cellular Biochemistry, Graduate School of Medical Sciences, Kyushu University, Fukuoka 812-8582, Japan.
Yamazaki
Soh
S
Eto-Kimura
Akiko
A
Takeshige
Koichiro
K
Muta
Tatsushi
T
D048868:Adaptor Proteins, Signal Transducing; D017398:Alternative Splicing; D000818:Animals; D019556:COS Cells; D002460:Cell Line; D019084:Fluorescent Antibody Technique, Indirect; D017353:Gene Deletion; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D006367:HeLa Cells; D006801:Humans; D020672:I-kappa B Proteins; D015151:Immunoblotting; D047468:Immunoprecipitation; D007249:Inflammation; D015850:Interleukin-6 / Q000378:metabolism; D051379:Mice; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000378:metabolism*; D041681:NIH 3T3 Cells; D009687:Nuclear Proteins / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D012190:Retroviridae / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014162:Transfection
D048868:Adaptor Proteins, Signal Transducing; D020672:I-kappa B Proteins; D015850:Interleukin-6; D016328:NF-kappa B; C417234:NFKBIZ protein, human; D009687:Nuclear Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins