The relationships between environmental factors and the genetic abnormalities that drive carcinogenesis are supported by experimental and epidemiologic evidence but their molecular basis has not been fully elucidated. At the genomic level, most human cancers display either chromosomal (CIN) or microsatellite (MIN) instability. The molecular mechanisms through which normal cells acquire these forms of instability are largely unknown. The arylamine 4-aminobiphenyl (4-ABP) is a tobacco smoke constituent, an environmental contaminant, and a well-established carcinogen in humans. Among others, bladder, lung, colon, and breast cancers have been associated with 4-ABP. We have investigated the effects of 4-ABP and N-methyl-N′-nitro-N-nitrosoguanidine (MNNG) on genetically stable colorectal (HCT116) and bladder (RT112) cancer cells. Cells were treated with carcinogens to generate resistant clones that were then subjected to genetic analysis to assess whether they displayed either CIN or MIN. We found that 50% to 60% of cells treated with 4-ABP developed CIN but none developed MIN as confirmed by their ability to gain and lose chromosomes. In contrast, all MNNG-treated clones (12/12) developed MIN but none developed CIN as shown by the microsatellite assay. The mismatch repair protein expression analysis suggests that the acquired mechanism of MIN resistance in the HCT116 MNNG-treated cells is associated with the reduction or the complete loss of MLH1 expression. By providing a mechanistic link between exposure to a tobacco constituent and the development of CIN, our results contribute to a better understanding of the origins of genetic instability, one of the remaining unsolved problems in cancer research.