The Drosophila protein Groucho is involved in the regulation of cell-determination events during insect neurogenesis and segmentation. A group of mammalian proteins, referred to as transducin-like Enhancer of split (TLE) 1 through 4, share with Groucho identical structures and molecular properties. The aim was to determine whether individual TLE proteins participate in the regulation of cell determination in mammals like their Drosophila counterpart. It is here reported that TLE family members are expressed in combinatorial ways during the in vitro differentiation of mouse P19 embryonic carcinoma cells (a model for neural determination) and rat CFK2 cells (a model for chondrocytic determination). TLE1 is up-regulated and TLE2 and TLE4 are down-regulated to different extents during early stages of differentiation. In contrast, later stages correlate with up-regulation of TLE2 and TLE4, and decreased expression of TLE1. Individual TLE proteins are also expressed in combinatorial as well as complementary patterns during the development of the cerebral cortex and spinal cord of mouse embryos. In particular, TLE1 is robustly expressed in both neural progenitor cells and postmitotic neurons of the outer layers of the cortical plate, whereas TLE4 expression marks preferentially postmitotic neurons of the inner layers. Taken together, these results strongly suggest non-redundant roles for individual TLE proteins during both cell-determination and cell-differentiation events.