CLONING OF NOVEL GENE WHOSE PRODUCT MEDIATES LIGAND-INDEPENDENT INDUCTION OF CYP1A1

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Our aim of this project is to identify and clone gene whose product mediates ligand-independent induction of CYP1A1 Previously, we showed that CYP1A1 expression can be induced by omeprazole (OP) in the human cell line HepG2,but not in the mouse cell line Hepa-1 cells. Now we show induction of CYP1A1 by α-naphthoflavone (αNF) in Hepa-1 cells. This induction was inhibited by the tyrosine kinase inhibitor herbimycin A, but not by the aromatic hydrocarbon (Ah)-receptor antagonist, suggesting the presence of a ligand-independent signal-transduction pathway in the mouse cell line too. We utilized the lack of CYP1A1 induction by OP in Hepa-1 cells to map a putative human gene OP-responsiveness in cell hybrids produced by fusion of Hepa-1 and HepG2 cells. OP-induced CYP1A1 expression was detected in four out of the 32 Hepa-1xHepG2 cell hybrids analyzed. To help identity the gene locus, a radiation-hybrid cell (E11 )was constructed Use of reverse-fluorescence in situ hybridization revealed that these five cell lines commonly retained human chromosome 10p. These results suggest that the human gene for OP-responsiveness is present on chromosome 10p13.