The LIM-homeodomain transcription factor LMX1B regulates expression of NF-kappa B target genes

LMX1B is a LIM-homeodomain transcription factor essential for development. Putative LMX1B target genes have been identified through mouse gene targeting studies, but their identity as direct LMX1B targets remains hypothetical. We describe here the first molecular characterization of LMX1B target gene regulation. Microarray analysis using a tetracycline-inducible LMX1B expression system in HeLa cells revealed that a subset of NF-κB target genes, including IL-6 and IL-8, are upregulated in LMX1B-expressing cells. Inhibition of NF-κB activity by short interfering RNA-mediated knock-down of p65 impairs, while activation of NF-κB activity by TNF-α synergizes induction of NF-κB target genes by LMX1B. Chromatin immunoprecipitation demonstrated that LMX1B binds to the proximal promoter of IL-6 and IL-8 in vivo, in the vicinity of the characterized κB site, and that LMX1B recruitment correlates with increased NF-κB DNA association. IL-6 promoter–reporter assays showed that the κB site and an adjacent putative LMX1B binding motif are both involved in LMX1B-mediated transcription. Expression of NF-κB target genes is affected in the kidney of Lmx1b −/− knock-out mice, thus supporting the biological relevance of our findings. Together, these data demonstrate for the first time that LMX1B directly regulates transcription of a subset of NF-κB target genes in cooperation with nuclear p50/p65 NF-κB.