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The glucocorticoid induction of alpha 1-acid glycoprotein (AGP) RNA in rat hepatoma cells is diminished by inhibiting protein synthesis. We now show that the AGP 5'-flanking region contains a DNA sequence (position -121 to -107), exhibiting a high degree of homology to the glucocorticoid regulatory element (GRE) consensus sequence ACAXXXTGTTCT, which serves to specifically bind purified rat glucocorticoid receptor in vitro. A 15 base pair oligonucleotide representing the AGP GRE confers glucocorticoid responsiveness on a heterologous promoter; such regulation is not diminished by concurrent inhibition of protein synthesis. However, inclusion of the AGP sequences immediately downstream of the AGP GRE (position -106 to -42) renders the hormonal induction sensitive to inhibition of protein synthesis. Furthermore, inclusion of these downstream sequences results in a more pronounced induction mediated by the AGP GRE. In vitro DNase-1 treatment using nuclear extracts prepared from HTC hepatoma cells generate footprints that indicate the presence of DNA-protein interactions spanning the region from -110 to -68 of the AGP gene. We propose that one or more labile factors acting within this domain, immediately downstream of the GRE, is required for efficient transcription at the AGP promoter.

In patients with cutaneous T-cell lymphomas (CTCL), soluble interleukin-2 receptor serum levels (sIL-2R) were determined by ELISA technique, and natural killer cell (NK) activity, by a 4-h chromium-51 release assay. Decrease of NK activity correlated with the augmentation of serum sIL-2R. After a 4-d stimulation with interleukin 2 CTCL patients' peripheral mononuclear cells (PMC) showed an increase of cytotoxic activity similar to that in healthy donors' PMC. Normal donors' PMC demonstrated a diminished IL-2-induced cytotoxic activity in 25% CTCL serum (sIL-2R of 3000, 7330, and 10700 U/ml, respectively) compared to control serum (sIL-2R of 400, 340, and 420 U/ml, respectively). IL-2-dependent proliferation of 2-d phytohemagglutinin (PHA) blasts was lower in CTCL serum than in control serum. sIL-2R was enriched from one CTCL patient's serum by IL-2 affinity chromatography. Transfection of the Tac gene into NIH/3T3 fibroblasts resulted in the production of a recombinant sIL-2R. The presence of enriched native or recombinant sIL-2R inhibited interleukin-2-dependent generation of cytotoxic activity and PHA blast proliferation. We suggest that elevated sIL-2R levels account for diminished NK activity by neutralizing interleukin 2 in CTCL patients.

Soluble interleukin-2 receptor (sIL-2R) alpha chain serum levels as determined by enzyme-linked immunosorbant assay (ELISA) are commonly used to monitor various inflammatory and neoplastic disorders associated with lymphocytic proliferation and activation. The in vivo structure of this soluble receptor species, however, is not characterized. We investigated sera with elevated sIL-2R serum levels of patients with histologically proven cutaneous T cell lymphoma (CTCL) and high-dose IL-2-treated melanoma patients and healthy donors. Purified recombinant IL-2R alpha and beta chain molecules, produced by transfection of NIH/3T3 fibroblasts and CHO cells, served as positive controls for purification and detection procedures. For selective enrichment of IL-2R molecules from supernatants and sera, affinity columns were prepared by coupling recombinant IL-2 or monoclonal antibodies against the alpha and the beta chain of the IL-2R complex to cyanogen bromide-activated Sepharose. Western blotting with affinity-purified fractions under reducing and nonreducing conditions revealed proteins that showed immunoreactivity for IL-2R alpha, beta, and gamma chain using several detection antibodies against these molecules. We conclude that the composition of sIL-2R in vivo is more complex than that of recombinant sIL-2R and can include all three IL-2R chains.

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