Cdx4 is a Cdx2 target gene

The products of the Cdx genes, Cdx1, Cdx2 and Cdx4, play multiple roles in early vertebrate development, and have been proposed to serve to relay signaling information from Wnt, RA and FGF pathways to orchestrate events related to anterior-posterior vertebral patterning and axial elongation. In addition, Cdx1 and Cdx2 have been reported to both autoregulate and to be subject to cross regulation by other family members. We have now found that Cdx4 expression is significantly down regulated in Cdx2(-/-) mutants suggesting previously unrecognized cross-regulatory interactions. Moreover, we have previously shown that Cdx4 is a direct target of the canonical Wnt signaling pathway, and that Cdx1 physically interacts with LEF/TCF members in an autoregulatory loop. We therefore investigated the means by which Cdx2 impacted on Cdx4 expression and assessed potential interaction between Cdx2 and canonical Wnt signaling on the Cdx4 promoter. We found that the Cdx4 promoter was regulated by Cdx2 in transient transfection assays. Electrophoretic mobility shift assays showed that Cdx2 bound to predicted Cdx response elements in the Cdx4 promoter which, when mutated, significantly reduced activity. Consistent with these data, chromatin immunoprecipitation assays from embryos demonstrated occupancy of the Cdx4 promoter by Cdx2 in vivo. However, we failed to observe an interaction between Cdx2 and components of the canonical Wnt signaling pathway. These findings suggest that, while both canonical Wnt and Cdx2 can regulate the activity of the Cdx4 promoter, they appear to operate through distinct mechanisms.