Structure, function, and regulation of the interleukin-2 receptor and identification of a novel immune activation gene

This chapter is divided into two sections, the first dealing with a novel immune activation gene, denoted Act-2. This gene encodes a secreted protein that may represent a new cytokine. The Act-2 protein shares significant homology with proteins in two related families of small secreted proteins. Act-2 is rapidly synthesized by activated T cells, B cells and monocytes. The second section deals with interleukin-2 receptors. These receptors are now known to be comprised of three distinct classes of receptors, formed by various combinations of two IL-2 binding proteins, the alpha and beta chains. The low-affinity receptors contain alpha, but not beta chains; the intermediate-affinity receptors contain beta, but not alpha chains, and the high-affinity receptors contain both alpha and beta chains. The beta chain appears to be tyrosine phosphorylated. We discuss evidence for the existence of another protein of relative molecular mass 100,000, which appears to be a subunit of at least the high-affinity receptor.     

Differential regulation of interleukin-6 receptor and gp130 gene expression in rat hepatocytes.

Interleukin-6 (IL-6) relays an important signal to hepatocytes during the early stages of an acute inflammatory response, causing an alteration in the expression of several major defense proteins. Additional regulation of this signal could occur either by altering the number of IL-6 receptors (IL-6-R) or of the signal transducing protein, gp130. We employed ribonuclease protection assays to measure the expression of IL-6-R and gp130 mRNA in primary rat hepatocytes in response to IL-6, interleukin-1, dexamethasone, and combinations thereof. Dexamethasone increases receptor mRNA levels 2.7-fold above controls but has no detectable effect on that of gp130. Such treatment increased surface expression of IL-6-R from 600 receptors per cell to greater than 6000, without a change in Kd (2.5-4.6 x 10(-10) M). In contrast to the stimulatory effect of the steroid signal, the inflammatory cytokines, individually and together, down-modulated both the mRNA and the cell surface expression of IL-6-R. These findings demonstrate for the first time that a sensitive control system exists between inflammatory mediators and IL-6-R.     

Nuclear factors interacting with an interleukin-6 responsive element of rat alpha 2-macroglobulin gene.

During acute inflammation, a group of liver-derived plasma proteins, acute phase proteins (APPs), increase in concentration. Interleukin-6 (IL-6) is responsible for this increase via the induction of APP gene expression. We have identified an IL-6 responsive cis-acting element (IL-6RE) of gene encoding a typical APP, rat alpha 2-macroglobulin (alpha 2M). The IL-6RE contains a sequence that is conserved among the 5'-flanking regions of various APP genes. Introduction of mutations into the conserved sequence revealed that the sequence, termed IL-6RE core, is a critical and essential component of IL6-RE. Nuclear factors binding to the IL-6RE core were identified in livers of normal and inflamed rats. Mobility shift pattern and DNase I footprinting profile indicated that the factors from normal and inflamed stages recognized the same sequence but were distinct from each other. These results suggested that the regulation of alpha 2M gene expression may involve mutually exclusive interaction of stage-specific trans-acting factors.     

Tissue specific hormonal regulation of the rat angiotensinogen gene expression

Angiotensinogen is the precursor of the most potent pressor substance, angiotensin. Angiotensinogen levels are increased in some forms of human hypertension. Its levels are modulated by various factors including glucocorticoids, estrogens, and prostaglandins. We have recently reported the isolation of a human angiotensinogen cDNA clone and provided evidence for the presence of its mRNA in rat liver, brain, and heart. In this communication we report the effect of dexamethasone and estradiol on angiotensinogen mRNA levels in rat liver, brain, and heart. Our results indicate that angiotensinogen levels are increased to different extents in these three tissues as a result of glucocorticoid or estrogen administration.     

Tissue-specific and hormonal regulation of the gene for rat prostatic steroid-binding protein in transgenic mice.

We investigated the tissue-specific and hormonal regulation of the gene for rat prostatic steroid-binding protein by introducing the C3(1) gene with 4-kilobase (kb) upstream and 2-kb downstream flanking sequences into transgenic mice. There was selective expression in the ventral prostate that was stimulated by testosterone, which indicated that the gene together with 6-kb flanking DNA contains the information required for prostate-specific and testosterone-regulated expression.     

Pneumadin in the rat ventral prostate and its hormonal regulation.

Pneumadin (PNM) is a decapeptide originally isolated from mammalian lungs, and exerts a potent antidiuretic action by stimulating arginine-vasopressin release. We have recently developed a sensitive and specific radioimmunoassay (RIA) for rat PNM and detected high concentrations of PNM--not only in the rat lungs, but also in the prostate. Hence, we investigated whether prostate PNM content is regulated by sex hormones. Male adult rats were orchidectomized or sham-operated and given a subcutaneous injection of testosterone or estradiol (40 and 5 mg/kg), respectively. The animals were decapitated one week after surgery, and their ventral prostates were promptly removed and weighed. PNM concentration and localization in the prostate were investigated by RIA and immunocytochemistry (ICC). Orchidectomy resulted in significant decreases in the prostate weight and PNM concentration, and testosterone administration prevented these effects. Estradiol administration to sham-operated rats caused prostate atrophy without changing PNM concentration. ICC localized PNM immunoreactivity (IR) exclusively in the epithelial cells of the ventral prostate. Orchidectomy markedly reduced PNM-IR concentration, while testosterone abolished this effect. Estradiol did not modify PNM-IR concentration in the atrophic prostate of sham-operated rats. We conclude that PNM content of rat prostate is dependent on the presence of adequate levels of circulating testosterone. The possibility that PNM plays a key role in the maintenance of the prostate growth is unlikely since estradiol-induced gland atrophy is not associated with any decrease in PNM concentration. The localization of PNM in the epithelial cells could suggest that this peptide may be involved in the regulation of some testosterone-dependent secretory functions of the rat prostate.     

Nucleotide sequence of the haptoglobin and haptoglobin-related gene pair. The haptoglobin-related gene contains a retrovirus-like element

Thirty-three kilobase pairs (kb) of human DNA containing the haptoglobin (Hp) and haptoglobin-related (Hpr) gene pair were cloned, and the nucleotide sequence of 21-kb DNA was determined. The two genes are closely linked, with Hpr being 2.2 kb downstream of Hp. Six hundred nucleotides of DNA occur between the two genes that are not found either 5' to the Hp gene or 3' to the Hpr gene. After the duplication event, the first intron of the Hpr gene acquired a 9-kb insert consisting mainly of a retrovirus-like element with a potential primer-binding site homologous to a mouse isoleucine tRNA. The element forms a repeated family in the human genome that I name RTVL-I (retrovirus-like element-isoleucine). In the coding region of the Hpr gene, there are no frameshift or nonsense mutations and its exon-intron splicing sites, 5' flanking and 3' flanking sequences do not show any obvious defects. There are 28 amino acid differences between the decoded amino acid sequences of the Hpr and Hp genes. Sixteen of these differences occur in the hpr beta chain, and all appear to be located on the surface of the molecule in places not thought to be involved in the hemoglobin binding function of haptoglobin. The structure of the Hpr gene suggests that the gene may be expressed and give rise to a functional product.