The caudal-related homeobox gene Cdx-2/3 is a critical "master" control gene in embryogenesis. Mice heterozygous for a null mutation in Cdx-2/3 exhibit multiple malfunctions including tail abnormalities, stunted growth, a homeotic shift in vertebrae, and the development of multiple intestinal adenomatous polyps, indicating that Cdx-2/3 is haplo-insufficient. In vitro studies have identified more than a half-dozen downstream target genes expressed in pancreatic and intestinal cells for this transcription factor. We have examined the transcriptional properties of the mouse Cdx-2/3 promoter. This promoter could be autoregulated in pancreatic and intestinal cells that express endogenous Cdx-2/3. In contrast, Cdx-2/3 transfection represses the Cdx-2/3 promoter in fibroblasts, which do not express endogenous Cdx-2/3. Since Cdx-2/3 activates proglucagon gene promoter in both pancreatic and intestinal cells and in fibroblasts, we suggest that some, yet to be identified, cell type-specific components are required for activating selected target gene promoters of Cdx-2/3, including the Cdx-2/3 promoter itself. Cdx-2/3 binds to the TATA box and another AT-rich motif, designated as DBS, within an evolutionarily conserved proximal element of the Cdx-2/3 promoter. The DBS motif is critical for the autoregulation, whereas the TATA box may act as an attenuating element for the autoregulatory loop. Finally, overexpression of Cdx-2/3 in a pancreatic cell line activated the expression of the endogenous Cdx-2/3. Taken together, our results indicate that the dose-dependent phenotype of Cdx-2/3 expression on its downstream targets in vivo could be regulated initially via a transcriptional network involving cell type-specific autoregulation of the Cdx-2/3 promoter.