IDENTIFICATION OF A CIS-ACTING ELEMENT THAT IS RESPONSIBLE FOR CADMIUM-MEDIATED INDUCTION OF THE HUMAN HEME OXYGENASE GENE

Heme oxygenase is an essential enzyme in heme catabolism and is induci ble by various environmental derangements such as cadmium. The activit y and mRNA levels of heme oxygenase were remarkably increased in HeLa cells by the treatment with cadmium. As a first step in studying the m olecular mechanisms of this induction, we performed transient expressi on assays in four human cell lines including HeLa to analyze the cadmi um-mediated inducibility of the fusion genes, containing the firefly l uciferase gene as a reporter under the human heme oxygenase gene promo ter. By determining the luciferase activity expressed in the transfect ed cells, we found the region between about 4.5 and 4 kilobase pairs u pstream from the transcriptional initiation site of the heme oxygenase gene which confers cadmium-mediated inducibility on the fusion gene. The region was then subjected to further functional analysis in HeLa c ells, which allowed us to localize the cadmium-responsive element to 2 0 base pairs. Gel mobility shift assays demonstrated that this 20-base pair element is specifically bound by nuclear protein(s) of HeLa cell s, the binding activities of which were however unchanged by the treat ment with cadmium. Using the synthetic cadmium-responsive elements con taining various base changes, we have identified a 10-base pair sequen ce, TGCTAGATTT, required for the cadmium-mediated inducibility and in vitro protein binding. We thus suggest that this binding protein(s) is involved in the cadmium-mediated activation of the heme oxygenase gen e. Incidentally, the consensus sequence of AP-1 binding site, TGAGTCA, is present downstream of this cadmium-responsive element. However, we provide evidence that AP-1 is not directly involved in the cadmium-me diated induction of the human heme oxygenase gene.