Cyclin D1 is required for transformation by activated Neu and is induced through an E2F-dependent signaling pathway

The neu (c-erbB-2) proto-oncogene encodes a tyrosine kinase receptor that i s overexpressed in 20 to 30% of human breast tumors. Herein, cyclin D1 prot ein levels were increased in mammary tumors induced by overexpression of wi ld-type Neu or activating mutants of Neu in transgenic mice and in MCF7 cel ls overexpressing transforming Neu, Analyses of 12 Neu mutants in MCF7 cell s indicated important roles for specific C-terminal autophosphorylation sit es and the extracellular domain in cyclin D1 promoter activation. Induction of cyclin D1 by NeuT involved Ras, Rac, Rho, extracellular signal-regulate d kinase, c-Jun N-terminal kinase, and p38, but not phosphatidylinositol 3- kinase, NeuT induction of the cyclin D1 promoter required the E2F and Spl D NA binding sites and was inhibited by dominant negative E2F-1 or DP-1, Neu- induced transformation was inhibited by a cyclin D1 antisense or dominant n egative E2F-1 construct in Rat-1 cells. Growth of NeuT-transformed mammary adenocarcinoma cells in nude mice was blocked by the cyclin D1 antisense co nstruct. These results demonstrate that E2F-1 mediates a Neu-signaling casc ade to cyclin D1 and identify cyclin D1 as a critical downstream target of neu-induced transformation.