Heme oxygenase (HO) is the rate-limiting enzyme in heme catabois and its activity is induced by many agents, including its substrate heme, heavy metals, UV radiation, and other injurious oxidant conditions. We examined the presence of several regulatory elements in the promoter region of the human HO-1 gene which could possibly account for its induction in response to diverse agents or influences. (11,12). Two HO isozymes, the products of two distinct genes, have been described (13,14). HO-1 is the inducible form which is ubiquitously distributed in mammalian tissues, whereas HO-2 is believed to be constitutively expressed, is not inducible by HO-1 inducers, and is present in tissues such as the brain and testis (14).One of the mechanisms by which hormones, growth factors, and other stimuli induce the expression of genes is by activating various transcription factors. This is a rapid process which frequently involves transcriptional or structural activation of the factor and allows its presence or transfer to the nucleus. These processes may be part of the mechanism by which various agents, including heme, increase HO expression and activity. Previous studies have shown the presence of AP-1-binding sequences and interleukin 6-, metal-, and heat-responsive elements in the HO-1 promoter region and have suggested the involvement of these nuclear factors in the regulation of several genes, including that encoding [13][14][15][16][17][18]. Erythropoietic cells are endowed with HO activity which is inducible by heme (6,7). We therefore used a human-derived erythroleukemic cell line, K562, to examine the presence of transcription factors which might be involved in heme-induced HO-1 expression and to determine whether binding sequences for these factors were present in the promoter region of the human HO-1 gene. tTo whom reprint requests should be addressed. MATERIALS AND METHODS 5987The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

these studies provides direct evidence that the inductive response of human HO to such injurious stimuli represents an important tissue adaptive mechanism for moderating the severity of cell damage produced by these blood components.

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An expression construct containing the cDNA encoding a modified aequorea green fluorescent protein (GFP) ligated to the 5'-end of the rat androgen receptor (AR) cDNA (GFP-AR) was used to study the intracellular dynamics of the receptor movement in living cells. In three different cell lines, ie. PC3, HeLa, and COS1, unliganded GFP-AR was seen mostly in the cytoplasm and rapidly (within 15-60 min) moved to the nuclear compartment after androgen treatment. Upon androgen withdrawal, the labeled AR migrated back to the cytoplasmic compartment and maintained its ability to reenter the nucleus on subsequent exposure to androgen. Under the condition of inhibited protein synthesis by cycloheximide (50 microg/ml), at least four rounds of receptor recycling after androgen treatment and withdrawal were recorded. Two nonandrogenic hormones, 17beta-estradiol and progesterone at higher concentrations (10(-7)/10(-6) M), were able to both transactivate the AR-responsive promoter and translocate the GFP-AR into the nucleus. Similarly, antiandrogenic ligands, cyproterone acetate and casodex, were also capable of translocating the cytoplasmic AR into the nucleus albeit at a slower rate than the androgen 5alpha-dihydrotestosterone (DHT). All AR ligands with transactivation potential, including the mixed agonist/antagonist cyproterone acetate, caused translocation of the GFP-AR into a subnuclear compartment indicated by its punctate intranuclear distribution. However, translocation caused by casodex, a pure antagonist, resulted in a homogeneous nuclear distribution. Subsequent exposure of the casodex-treated cell to DHT rapidly (15-30 min) altered the homogeneous to punctate distribution of the already translocated nuclear AR. When transported into the nucleus either by casodex or by DHT, GFP-AR was resistant to 2 M NaCl extraction, indicating that the homogeneously distributed AR is also associated with the nuclear matrix. Taken together, these results demonstrate that AR requires ligand activation for its nuclear translocation where occupancy by only agonists and partial agonists can direct it to a potentially functional subnuclear location and that one receptor molecule can undertake multiple rounds of hormonal signaling; this indicates that ligand dissociation/inactivation rather than receptor degradation may play a critical role in terminating hormone action.