MEOX2 regulates nuclear factor-κB activity in vascular endothelial cells through interactions with p65 and IκBβ

Tumours secrete proangiogenic factors to induce the ingrowth of blood vessels, the end targets of which are vascular endothelial cells (ECs). The MEOX2 homeoprotein inhibits nuclear factor-kappa B (NF-kappa B) signalling and EC activation in response to serum and proangiogenic factors. We hypothesize that MEOX2 interacts with components of this pathway in vascular ECs to modulate NF-kappa B activity and EC activation and that these interactions depend upon specific domains within the MEOX2 protein. To test our hypothesis, we transduced ECs with MEOX2 expression constructs. MEOX2 protein localized to the nuclear fraction, as did I kappa B beta and p65. By co-immunoprecipitation, MEOX2 bound to both p65 and I kappa B beta. Immunofluorescence demonstrated that MEOX2 colocalizes in the nucleus with both p65 and I kappa B beta and that this colocalization requires the MEOX2 homeodomain and N-terminal domain. Finally, promoter assays revealed that MEOX2 expression has a biphasic effect on NF-kappa B-dependent promoters. At low levels, MEOX2 stimulates NF-kappa B activity, whereas at high levels, it represses, effects that also depend upon the homeodomain and the N-terminal domain. Our results represent the first report of an interaction between a homeobox protein and I kappa B beta and suggest that MEOX2 modulates the activity of the RelA complex through direct interaction with its components. These observations implicate MEOX2 as a potentially important regulatory gene inhibiting not only the angiogenic response of ECs to proangiogenic factors, but also their response to chronic inflammatory stimulation that normally activates NF-kappa B, suggesting MEOX2 as a possible molecular target for the therapy of angiogenesis-dependent diseases such as cancer.