Saatcioglu, Fahri
Multiple DNA-protein interactions at the aryl hydrocarbon (Ah)- responsive enhancer of the CYP1A1 gene.

Order No. 9025204

The polycyclic aromatic hydrocarbon-inducible cytochrome P450IA1 (CYP1A1) gene is capable of both detoxifying polycyclic aromatic compounds, such as tetrachlorodibenzo-p- dioxin (TCDD), 3-methylcholanthrene and beta-napthoflavone, and converting some compounds to highly active carcinogens and mutagens. Both transcriptional and post-transcriptional mechanisms have been shown to be important in the regulation of the CYP1A1 gene. Previously, two xenobiotic response elements, XRE1 and XRE2, both of which can activate transcription of the CYP1A1 gene, were identified. We present evidence in the following chapters that three distinct factors, XF-A1, XF-A2, and XF-B, which has been identified as the Ah receptor, interact specifically with the XRE sequence motif at the PAH-responsive enhancer of the CYP1A1 gene. XF-A1 and XF-A2, which bind to the XRE motif, have not been described previously. Our footprinting analysis shows that, although XF-A1 and XF-A2, and the Ah receptor all bind to the same 15 base pair sequence domain, they interact with distinctly different nucleotides. Our analysis indicates that XF-A1 and XF-A2, and the Ah receptor are the products of different genes, that their expression is regulated differently, and that these two factors require different cofactors. Comparison of the binding activities of the Ah receptor, XF-A1 and XF-A2 for XRE1 and XRE2 reveal substantial differences in affinity. In addition, there are differences in the interaction of the Ah receptor with XRE1 and XRE2.

In particular, the following points can be made regarding these three factors: (1) XF-A1, XF-A2, and the Ah receptor are probably different gene products, since mutant cell lines defective in the Ah receptor have normal levels of XF-A1 and XF-A2 activity. (2) XF-A1 and XF-A2 activities are constitutively expressed in the nucleas whereas Ah receptor- related DNA-binding activity appears in the nucleus only after treatment with polycyclic compounds. (3) Zn2+ is an essential cofactor for the Ah receptor, but not for XF-A1 or XF-A2, consistent with the earlier hypothesis that the Ah receptor is a member of the steroid and thyroid hormone receptor super- family. (4) The affinities of the Ah receptor, XF-A1 and XF-A2 for XRE2 are 15- and 50- fold weaker, respectively, than for XRE1, which suggests that the two XREs may differ in their contribution to CYP1A1 gene transcription. (5) In contrast to previous studies in Hepa1 cells, a single copy of either XRE has no discernible transcriptional activity in primary cultures of rat hepatocytes, but a 115-bp region containing both XREs has enhancer activity in response to both TCDD and cycloheximide. [Abstract shortened.] Source: DAI, 51, no. 08B, (1990): 3696


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