We have explored this hypothesis and confirmed that pRb is required to achieve normal levels of detoxification enzymes and, furthermore, that depletion of activator E2Fs seems to play a similar role. Our results are indicative that both pRb and E2F1-2-3 act as activators of the detoxification system and have important implications for understanding tumor progression and tumor treatment. The E2F factors regulate multiple genes essential to progress through S phase, and binding of E2Fs by the retinoblastoma related proteins effectively blocks S-phase progression and suppresses cell proliferation. Associations between pRb proteins and E2Fs are normally regulated by cyclin-dependent phosphorylation, ensuring a smooth transition through the cell cycle. Exogenous factors, including viral oncoproteins like SV40 T antigen, can disrupt the pRb/E2F complexes, resulting in upregulation of E2F target genes, uncontrolled proliferation and/ or tumorigenesis both in vivo and in vitro. An LXCXE peptide motif in TAg is Senegenin necessary to bind the pRb proteins and to induce cell proliferation. In addition to the induction of ectopic proliferation, expression of TAg results in the mRNA downregulation of detoxification components from Phases I, II and III in intestinal enterocytes. This effect requires an intact LXCXE motif in TAg, suggesting that pRb might control the transcription of detoxification components. To explore this possibility, we examined the RNA levels of different members of Phases I, II and III in genetic background devoid of pRb. Conventional inactivation of pRb in mice results in numerous alterations and null embryos die around E13.5, hampering the study of possible pRb effects prior to birth. Nevertheless, pRb-deficient embryos supplied with a wild-type placenta are viable, but die soon after birth. Using such mice allowed us to analyze the effects of pRb ablation in tissues from embryos and newborn mice. At E12.5, the earliest point analyzed, the Isochlorogenic-acid-C endogenous expression of detoxification components is absent or barely detectable even by in situ hybridization. However, the transcription levels of Cyp2d10 in livers of control embryos generated from females treated with PCN an inducer of the detoxification pathway, are substantially increased, while are not detectable in RBKO littermates. Furthermore, removal of pRb in E18.5 liver samples, at which point endogenous and inducible levels of several detoxification components are detectable, results in overall reduction of both endogenous and PCN-induced levels of Phases I, II and III transcripts, with the exception of Cyp1b1. We analyzed several tissues in newborn mice, and confirmed that removal of pRb results in a reduction of endogenous Cyp levels in newborn livers and intestines, while other transcripts used as control remain unaltered. This effect was confirmed in all the tissues expressing Cyps at that specific developmental stage, including liver, intestine and lungs. Other tissues analyzed from E18.5 to newborn mice, including brain, heart and spleen, did not show detectable RNA levels of members of the detoxification pathway with or without chemical challenge. Thus, it appears that pRb is required for constitutive expression and induction of the Cyp pathway. To determine if the observed changes in mRNA abundance are reflected in protein levels, we analyzed the levels of Cyp3A proteins, one of the most important subfamilies involved in the metabolism of xenobiotics, in intestinal and hepatic samples. All samples tested, including newborn intestines and livers as well as murine adult villi, showed a considerable decrease in Cyp3A proteins in the absence of pRb. Furthermore, removal of either pRb or E2F1-2-3 leads to decreased Cyp3A protein levels in the intestinal epithelium of adult mice. Thus, transcriptional downregulation of Cyp3A correlates with reduction in protein levels in intestinal and hepatic tissues.