Citation

The nuclear protein IB, comprising the N-terminal trans-activation domain and the C-terminal ankyrin repeat (ANK) domain composed of seven ANK motifs, activates transcription of a subset of NF-B-dependent innate immune genes such as Lcn2 encoding the antibacterial protein lipocalin-2. Lcn2 activation requires formation of a complex containing IB and NF-B p50, a transcription factor that harbors the DNA-binding Rel-homology region but lacks a trans-activation domain, on the promoter with the canonical NF-B-binding site (B site) and its downstream cytosine-rich element. Here we show that IB productively interacts with p50 via Asp-451 in the N-terminus of ANK1, a residue that is evolutionarily conserved among IB and the related nuclear IB proteins Bcl-3 and IBNS; threonine substitution for Asp-451 abrogates direct association with the B-site-binding protein p50, complex formation with the Lcn2 promoter DNA, and activation of Lcn2 transcription. The basic residues Lys-717 and Lys-719 in the C-terminal region of ANK7 contribute to IB binding to the Lcn2 promoter probably via interaction with the cytosine-rich element required for Lcn2 activation; glutamate substitution for both lysines results in a loss of transcriptionally-active complex formation without affecting direct contact of IB with p50. Both termini of the ANK domain in Bcl-3 and IBNS function in a manner similar to that of IB to interact with promoter DNA, indicating a common mechanism in which the nuclear IBs form a regulatory complex with NF-B and promoter DNA via the invariant aspartate in ANK1 and the conserved basic residues in ANK7.