Several studies have reported a correlation between expression of the estrogen receptor (ER) and aryl hydrocarbon (Ah) responsiveness in human breast cancer cell lines. MDA-MB-231 cells are ER-negative and Ah-nonresponsive; however, initial studies showed that 2,3,7,8-tetrachlorodibenzo-p-dioxin induced CYP1A1 mRNA levels (5.8-fold) and chloramphenicol acetyltransferase activity (2.6-fold) in high passage (Hp, >50 passages) cells transiently transfected with an Ah-responsive plasmid. In contrast, no induction responses were observed in low passage (Lp, <20 passages) cells. The Ah responsiveness of Hp compared to Lp MDA-MB-231 cells was associated with a >2-fold increased expression of the Ah receptor in Hp cells. Further analysis revealed that the apparent molecular weight of the Ah receptor mRNA transcript and immunoreactive protein were comparable in Lp MDA-MB-231 and Ah-responsive human HepG2 cells. In contrast, RT-PCR analysis of the Ah receptor nuclear translocator (Arnt) protein showed that HepG2 cells expressed the expected 2.6-kb transcript, whereas a 1.3-kb transcript was the major product in MDA-MB-231 cells. Western blot analysis confirmed that HepG2 cells primarily expressed a 97-kDa wild-type form of Arnt, whereas a dominant 36-kDa variant was expressed in MDA-MB-231 cells. Complete sequence analysis of the variant form of Arnt revealed a major deletion of the C-terminal region of the protein (aa 330 to 789). Like HepG2 cells, the wild-type 2.6-kb transcript was detected in ER-positive (Ah-responsive) MCF-7 cells, whereas the low-molecular-weight variant Arnt was dominant in ER-negative MDA-MB-231, MDA-MB-435, and Adriamycin-resistant MCF-7 cells. These results suggest that expression of this protein may be useful as a prognostic factor in breast cancer.