The Drosophila hindgut develops three morphologically distinct regions along its anteroposterior axis: small intestine, large intestine and rectum. Single-cell rings of 'boundary cells' delimit the large intestine from the small intestine at the anterior, and the rectum at the posterior. The large intestine also forms distinct dorsal and ventral regions; these are separated by two single-cell rows of boundary cells. Boundary cells are distinguished by their elongated morphology, high level of both apical and cytoplasmic Crb protein, and gene expression program. During embryogenesis, the boundary cell rows arise at the juxtaposition of a domain of Engrailed (En)- plus Invected (Inv)-expressing cells with a domain of Delta (Dl)-expressing cells. Analysis of loss-of-function and ectopic expression phenotypes shows that the domain of Dl-expressing cells is defined by En/Inv repression. Further, Notch pathway signaling, specifically the juxtaposition of Dl-expressing and Dl-non-expressing cells, is required to specify the rows of boundary cells. This Notch-induced cell specification is distinguished by the fact that it does not appear to utilize the ligand Serrate and the modulator Fringe.