In this study, we describe the isolation and characterization of Foxp4, a new member of the Foxp subfamily of winged-helix transcription factors. The full-length mouse Foxp4 cDNA encodes a 685-amino-acid protein that is similar to Foxp1 and Foxp2. Foxp4 gene expression is observed primarily in pulmonary, neural, and gut tissues during embryonic development. To compare the protein expression patterns of Foxp4 to Foxp1 and Foxp2, specific polyclonal antisera to each of these proteins was used in immunohistochemical analysis of mouse embryonic tissues. All three proteins are expressed in lung epithelium with Foxp1 and Foxp4 expressed in both proximal and distal airway epithelium while Foxp2 is expressed primarily in distal epithelium. Foxp1 protein expression is also observed in the mesenchyme and vascular endothelial cells of the lung. At embryonic day 12.5, Foxp1 and Foxp2 are expressed in both the mucosal and epithelial layers of the intestine. However, Foxp2 is expressed only in the outer mucosal layer of the intestine and stomach later in development. Finally, Foxp4 is expressed exclusively in the epithelial cells of the developing intestine, where, in late development, it is expressed in a gradient along the longitudinal axis of the villi.