Identification of a novel inhibitor of nuclear factor-kappaB, RelA-associated inhibitor.
Here we report the identification and characterization of a novel protein, RelA-associated inhibitor (RAI), that binds to the NF-kappaB subunit p65 (RelA) and inhibits its transcriptional activity. RAI gene was isolated in a yeast two-hybrid screen using the central region of p65 as bait. We confirmed the physical interaction in vitro using recombinant proteins as well as in vivo by immunoprecipitation/Western blot assay. RAI gene encodes a protein with homology to the C-terminal region of 53BP2 containing four consecutive ankyrin repeats and an Src homology 3 domain. RAI mRNA was preferentially expressed in human heart, placenta, and prostate. Despite its similarity to 53BP2, RAI did not interact with p53 in a yeast two-hybrid assay. RAI inhibited the action of NF-kappaB p65 but not that of p53 in transient luciferase gene expression assays. Similarly, RAI inhibited the endogenous NF-kappaB activity induced by tumor necrosis factor-alpha. RAI specifically inhibited the DNA binding activity of p65 when co-transfected in 293 cells. RAI protein appeared to be located in the nucleus and colocalized with NF-kappaB p65 that was activated by TNF-alpha. These observations indicate that RAI is another inhibitor of NF-kappaB in addition to IkappaB proteins and may confer an alternative mechanism of regulation.
Yang
J P
JP
Department of Molecular Genetics, Nagoya City University Medical School, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601, Japan.
Hori
M
M
Sanda
T
T
Okamoto
T
T
D000595:Amino Acid Sequence; D017487:Ankyrins / Q000737:chemistry; D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics*; D002460:Cell Line; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000037:antagonists & inhibitors* / Q000235:genetics*; D009687:Nuclear Proteins / Q000737:chemistry / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D012097:Repressor Proteins; D016415:Sequence Alignment; D051996:Transcription Factor RelA; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors / Q000494:pharmacology; D015003:Yeasts / Q000235:genetics
D017487:Ankyrins; D051017:Apoptosis Regulatory Proteins; D002352:Carrier Proteins; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; D016328:NF-kappa B; D009687:Nuclear Proteins; C119612:PPP1R13L protein, human; D012333:RNA, Messenger; D012097:Repressor Proteins; C090468:TP53BP2 protein, human; D051996:Transcription Factor RelA; D014409:Tumor Necrosis Factor-alpha