Groucho is a protein implicated in Notch signaling and involved in segmentation and neural development in Drosophila. Groucho forms transcription complexes with the basic helix-loop-helix proteins encoded by the hairy/Enhancer of split ("hairy-like") gene family. These interactions are mediated by the carboxyl-terminal WRPW motif of Hairy-like proteins. We are interested in determining whether Groucho and its mammalian homologues, the TLE proteins, perform conserved functions. We show that TLE1 interacts with HES-1, a murine homologue of Drosophila Hairy-like proteins, both in the yeast two-hybrid assay and in an interaction assay based on glutathione S-transferase fusion proteins. These results show that Groucho/TLE proteins and Hairy-like/HES proteins are involved in similar interactions in Drosophila and mammals and further suggest that these proteins perform conserved cellular functions.