CYP1A1 and CYP1B1 metabolically activate many polycyclic aromatic hydrocarbons (PAHs), including benzo[a]pyrene, to reactive intermediates associated with toxicity, mutagenesis, and carcinogenesis. Paradoxically, however, Cyp1a1^-/- knockout mice are more sensitive to oral benzo[a]pyrene exposure, compared with wild-type Cyp1a1^+/+ mice (Mol Pharmacol 65:1225, 2004). To further investigate the mechanism for this enhanced sensitivity, Cyp1a1^-/-, Cyp1a2^-/-, and Cyp1b1^-/- single-knockout, Cyp1a1/1b1^-/- and Cyp1a2/1b1^-/- double-knockout, and Cyp1^+/+ wild-type mice were analyzed. After administration of oral benzo[a]pyrene (125 mg/kg/day) for 18 days, Cyp1a1^-/- mice showed marked wasting, immunosuppression, and bone marrow hypocellularity, whereas the other five genotypes did not. After 5 days of feeding, steady-state blood levels of benzo[a]pyrene were ∼25 and ∼75 times higher in Cyp1a1^-/- and Cyp1a1/1b1^-/- mice, respectively, than in wild-type mice. Benzo[a]pyrene-DNA adduct levels were highest in liver, spleen, and marrow of Cyp1a1^-/- and Cyp1a1/1b1^-/- mice. Many lines of convergent data obtained with oral benzo[a]pyrene dosing suggest that: 1) inducible CYP1A1, probably in both intestine and liver, is most important in detoxication; 2) CYP1B1 in spleen and marrow is responsible for metabolic activation of benzo[a]pyrene, which results in immune damage in the absence of CYP1A1; 3) both thymus atrophy and hepatocyte hypertrophy are independent of CYP1B1 metabolism but rather may reflect long-term activation of the aryl hydrocarbon receptor; and 4) the magnitude of immune damage in Cyp1a1^-/- and Cyp1a1/1b1^-/- mice is independent of plasma benzo[a]pyrene and total-body burden and clearance. Thus, a balance between tissue-specific expression of the CYP1A1 and CYP1B1 enzymes governs sensitivity of benzo[a]pyrene toxicity and, possibly, carcinogenicity.