Inhibition of nuclear factor-κB-mediated transcription by association with the amino-terminal enhancer of split, a Groucho-related protein lacking WD40 repeats

The amino-terminal enhancer of split (AES) encodes a 197-amino acid protein that is homologous to the NH2-terminal domain of the Drosophila Groucho protein but lacks COOH-terminal WD40 repeats. Although the Drosophila Groucho protein and its mammalian homologs, transducin-like enhancer of split proteins, are known to act as non-DNA binding corepressors, the role of the AES protein remains unclarified. Using the yeast two-hybrid system, we have identified the protein-protein interaction between AES and the p65 (Re1A) subunit of the transcription factor nuclear factor kappa B (NF-kappa B), which activates various target genes involved in inflammation, apoptosis, and embryonic development. The interaction between AES and p65 was confirmed by in vitro glutathione S-transferase pull down assay and by in vivo co-immunoprecipitation study. In transient transfection assays, AES repressed p65-driven gene expression. AES also inhibited NF-kappa B-dependent gene expression induced by tumor necrosis factor alpha, interleukin-1 beta, and mitogen-activated protein kinase/extracellular signal-regulated kinase kinase kinase 1, which is an upstream kinase for NF-kappa B activation. These data indicate that AES acts as a corepressor for NF-kappa B and suggest that AES may play a pivotal role in the regulation of NF-kappa B target genes.