IDENTIFICATION OF BINDING-SITES FOR TRANSCRIPTION FACTORS NF-KAPPA-B AND AP-2 IN THE PROMOTER REGION OF THE HUMAN HEME OXYGENASE-1 GENE

Heme oxygenase (HO) is the rate-limiting enzyme in heme catabolism and its activity is induced by many agents, including its substrate heme, heavy metals, UV radiation, and other injurious oxidant conditions. W e examined the presence of several regulatory elements in the promoter region of the human HO-1 gene which could possibly account for its in duction in response to diverse agents or influences. Heme treatment in creased both HO activity and HO-1 mRNA in the human erythroleukemic ce ll line K562. Electrophoretic mobility-shift assays of nuclear protein extracts from heme-treated and control cells with specific oligonucle otide probes containing binding sites for known transcription factors, including AP-1, AP-2, Sp1, NF-kappa B, CTF/NF1, TFIID, OKT1, and CREB , and oligonucleotides containing serum-, metal-, and glucocorticoid-r esponsive elements demonstrated a specific and marked increase in the NF-kappa B and AP-2 transcription factors and, to a lesser extent, an increase in AP-1. No significant increase in other transcription facto rs over the control, untreated cells was observed. DNase I footprint a ssays using purified transcription factors revealed the presence of NF -kappa B and AP-2 binding sites in the proximal part of the promoter r egion of the human HO-1 gene. Moreover, nucleotide sequence analysis o f the HO-1 promoter region showed that the protected regions encompass ed NF-kappa B and AP-2 consensus binding sites. The presence of regula tory sequences for the binding of transcription factors such as NF-kap pa B and AP-2, whose activation is associated with the immediate respo nse of the cell to an injury, may be an indication of the important ro le which HO-1 may play in defense mechanisms against tissue injury.