Loss of cell cycle regulation in mammary epithelium results in impaired mammary gland development and neoplasia. We investigated the consequences of the absence of pRb in mammary epithelial cells during normal development and in mice that express an oncogene in the mammary epithelium. Since pRb-deficiency results in embryonic lethality, we transplanted pRb-null mammary anlagen into wild hosts. pRb-deficient mammary epithelia were capable of functional differentiation in term animals and they regenerated a differentiated gland even after multiple pregnancies. In serial transplantations no significant differences were found in outgrowth of pRb-deficient and wild type epithelia indicating that the absence of pRb does not lead to transformation. Likewise the effect of a TGFbeta1 transgene was not altered in the absence of pRb. The susceptibility of mammary epithelium to form tumors was assessed in three different models. No differences in tumor incidence were found between wild type and Rb +/- WAP-int3, MMTV-PyMT transgenic and Brcal-/- epithelia. These results demonstrate that the absence of pRb does not affect normal mammary gland development and tumorigenesis in three different mouse models investigated and suggest that loss of more than one member of the pRb pathway is required to induce mammary tumors.