Ebp1, an ErbB-3 binding protein, interacts with Rb and affects Rb transcriptional regulation

Ebp1, an ErbB-3 binding protein, inhibits the proliferation and induces the differentiation of human breast cancer cells. The mechanisms of these effe cts are unknown. Rb, the product of the retinoblastoma gene, is an importan t modulator of cell cycle progression and cellular differentiation. We repo rt that Rb is a binding target for Ebp1. Ebp1 was localized to both the nuc leus and the cytoplasm of logarithmically growing AU565 breast cancer cells and HeLa cells as determined by confocal immunofluorescent microscopy. Ebp 1 was present in Rb immunoprecipitates derived from AU565 breast cancer cel ls. GST-Rb also bound endogenous Ebp1. Using GST-Ebp1 constructs, we determ ined that the 72 C-terminal amino acids of Ebp1 were sufficient to bind Rb. Dephosphorylation of Ebp1 enhanced the interaction of Ebp1 with Rb. The ov erexpression of Ebp1 in MCF-7 and AU565 (Rb+) cells inhibited the activity of the E2F1 regulated cyclin-E promoter. Ebp1 bound E2F1 indirectly via Rb in lysates of MCF-7 cells. The interaction of Ebp1 with Rb may prove to be an important mechanism of Ebp1 induced changes in cell proliferation and di fferentiation. (C) 2001 Wiley-Liss. Inc.