Steroid receptor binding factor (RBF) was originally isolated from avian oviduct nuclear matrix. When bound to avian genomic DNA, RBF generates saturable high-affinity binding sites for the avian progesterone receptor (PR). Recent studies have shown that RBF binds to a 54 bp element in the 5'-flanking region of the progesterone-regulated avian c-myc gene, and nuclear matrix-like attachment sites flank the RBF element [Lauber et al. (1997) J. Biol. Chem. 272, 24657-24665]. In this paper, electrophoretic mobility shift assays (EMSAs) and S1 nuclease treatment are used to demonstrate that the RBF-maltose binding protei (MBP) fusion protein binds to single-stranded DNA of its element. Only the N-terminal domain of RBF binds the RBF DNA element as demonstrated by southwestern blot analyses, and by competition EMSAs between RBF-MBP and the N-terminal domain. Mass spectrometric analysis of the C-terminal domain of RBF demonstrates its potential to form noncovalent protein-protein interactions via a potential leucine-isoleucine zipperlike structure, suggesting a homo- and/or possible heterodimer structure in solution. These data support that the nuclear matrix binding site (acceptor site) for PR in the c-myc gene promoter is composed of RBF dimers bound to a specific single-stranded DNA element. The dimers of RBF are generated by C-terminal leucine zipper and the DNA binding occurs at the N-terminal parallel beta-sheet DNA binding motif. This complex is flanked by nuclear matrix attachment sites.

During the past few years, our understanding of nuclear receptor action has dramatically improved as a result of the identification and functional analysis of co-regulators such as factors involved in chromatin remodeling, transcription intermediary factors (co-repressors and co-activators), and direct interactions with the basal transcriptional machinery. Furthermore, the elucidation of the crystal structures of the empty ligand-binding domains of the nuclear receptor and of complexes formed by the nuclear receptor's ligand-binding domain bound to agonists and antagonists has contributed significantly to our understanding of the early events of nuclear receptor action. However, the picture of hormone- and hormone receptor-mediated mechanisms of gene regulation remain incomplete and extremely complicated when one also considers the "nontraditional" interactions of hormone-activated nuclear receptors, for example, interactions between the activated steroid receptors and components of the chromatin/nuclear matrix; and finally the nongenomic effects that steroid hormones can exhibit with other signaling pathways. In this prospectus on steroid receptors, we discuss the implications of various steroid hormone and nuclear receptor interactions and potential future directions of investigation.

The receptor-binding factor (RBF) for the avian oviduct progesterone (Pg) receptor (PR) has previously been shown to be a unique 10-kDa nuclear matrix protein that generates high affinity PR-binding sites on avian DNA. This paper describes the use of Southwestern blot and DNA gel shift analyses with RBF protein to identify a minimal 54-base pair RBF-binding element in the matrix-associated region (MAR) of the Pg-regulated c-myc gene promoter, This element contains a 5'-GC-rich domain and a 3'-AT-rich domain, the latter of which has a homopurine/homopyrimidine structure. The gel shift assays required the generation of an RBF-maltose fusion protein (RBF-MBP), which specifically binds this element and is supershifted when the anti-RBF polyclonal antibody is added. Computer analysis of the full-length amino acid sequence for RBF predicts a DNA-binding motif involving a beta-sheet structure at the N-terminal domain. Southern blot analyses using nuclear matrix DNA suggests that there are dual MAR sites in the c-myc promoter, which flank an intervening domain containing the RBF element. The co-transfection of this MAR sequence, containing the RBF element and cloned into a luciferase reporter vector, together with an RBF expression vector construct, into steroid treated human MCF-7 cells, results in a decrease of the c-myc promoter activity relative to control transfections containing only the parent vector of the RBF expression construct. These data suggest that a unique chromatin/nuclear matrix structure, composed of the RBF-DNA element complex which is flanked by nuclear matrix attachment sites, serves to bind the PR and repress the c-myc promoter.

We have previously proposed a novel mechanism for the coupled regulation of glucocorticoid receptor (GR) and c-jun transcription in triamcinolone acetonide (TA)-treated AtT-20 cells. This involved transcriptional interference of AP-1 (Fos/Jun)-driven gene transcription by the formation of inactive GR/Jun heterodimers. To further elucidate the molecular mechanism for GR autoregulation. the expression of GR and c-jun mRNA and protein levels were examined in both mouse L929 fibroblast cells and human CEM-C7 acute lymphoblastic leukemia cells. A rapid down-regulation of both GR and c-jun mRNA and protein levels occurs in TA-treated L929 cells. All-trans-retinoic acid (RA) treatment of Jun-deficient, mouse F9, teratocarcinoma cells causes the induction of c-jun expression. The increased expression of both c-jun mRNA and protein is accompanied by the induction of GR expression. These data further suggest that functional cJun is needed for the expression of the GR and c-jun genes in F9 cells. CEM-C7 cells undergo apoptosis after exposure to glucocorticoids. There is a parallel up-regulation of GR and c-jun mRNA levels in TA-treated CEM-C7 cells. This is accompanied by a concomitant increase in GR and cJun protein levels. Dose-response analyses reveal the expected coordinate regulation of both GR and c-jun mRNA and protein in L929 cells (decreasing) and in CEM-C7 cells (increasing). However, similar to 20-fold less TA is required for the inhibition of GR and c-jun expression as compared to that required for the stimulation of these two genes. These data demonstrate that the coordinate regulation of GR and c-jun gene expression is dose-dependent and cell type-specific. These results, along with previously reported data, suggest that GR complex formation with itself or with another transcription factor is important for the coordinate up- and down-regulation, respectively, of the GR and c-jun genes.

Autologous regulation of steroid receptors by their cognate ligands has been demonstrated for a number of nuclear receptor family members. To determine the molecular mechanism for glucocorticoid receptor (GR) autoregulation, the expression of glucocorticoid receptor mRNA and protein levels were examined in the mouse AtT-20 pituitary tumor cell line. The expression of c-jun and c-fos mRNA and protein was also examined in the same cell extracts. A rapid down-regulation of the GR protein was observed after treatment with the glucocorticoid analog, triamcinolone acetonide (TA). An oscillatory, parallel regulation of both GR and c-jun mRNA levels occurred. In contrast, POMC mRNA levels remained at a stable, low level during chronic TA treatment. Dose-response analyses also revealed a coordinate down-regulation of GR and c-jun (but not POMC or c-fos) mRNA levels. FOS protein levels were unaffected by TA treatment. Surprisingly, JUN protein levels were increased by TA, even when the c-jun mRNA levels were decreasing. Perhaps a derepression of c-jun mRNA translation occurs after TA treatment, and this may be due to GR/JUN heteromer formation interfering with JUN repression of c-jun mRNA translation. The effect of TA on GR and c-jun gene expression was a primary effect, as it occurred rapidly and was not inhibited by cycloheximide (CHX). Nuclear run-on transcription assays revealed a rapid (15 min) down-regulation in both GR and c-jun gene transcription rates, while POMC gene transcription was unaffected at this early time. Treatment of AtT-20 cells with all-trans retinoic acid gave different kinetics for GR and c-jun mRNA regulation than obtained with TA; however, the GR and c-jun mRNA levels were still coordinately regulated after retinoic acid treatment. Based upon these data, the promoter structures of the GR and c-jun genes, and previously published results, a novel mechanism for the coupled regulation of GR and c-jun transcription, via a direct transcriptional interference with AP-1 (FOS/JUN) activity, is proposed.

The major impetus for health care reform clearly seems to be the desire to contain the cost of health care. One of the major cost‐containment methods proposed is capitation. Capitation poses specific problems when applied to mental health services delivery. This article discusses several of the major issues pertaining to the utility of capitation in mental health, discusses on‐going capitation efforts, and provides an evaluation of capitation programs. Future directions for research on capitation in mental health are then suggested.

With increasing age, the thoracic aorta shows progressive fibroplastic intimal thickening, which is thought to be pre-atheromatous. A similar progressive intimal thickening in the renal cortical arteries is the distinguishing feature of the nephrosclerosis which underlies essential hypertension. Therefore, the earliest detectable youthful precursors of atherosclerosis and hypertension show strong morphological resemblances to each other. In this study, close statistical associations have been shown between the two types of arterial intimal fibroplasia. Both conditions show similar sigmoid growth curves from ages 6 to 70 years, thereby generating correlations across age groups of r = 0.99 in New Orleans and r = 0.95 in Mexico City. Specimens gathered in New Orleans were found to have about 1.4 times greater arterial intimal thickening than specimens from Mexico City, and this excess was seen at all ages in both the aortas and the renal cortical arteries. It seems likely that intimal fibroplasia of arteries is reflecting similar biological principles at all levels of the vascular tree. Whatever etiological factors vary between New Orleans and Mexico City, those factors appear to act directly at a tissue level to promote the early precursors of atherosclerosis and of the nephrosclerosis that underlies hypertension.

Performance contracting in Colorado continues to change based on changing system goals and finite resources. An earlier 1986–87 model along with its unresolved issues is linked to recent developments. Currently the areas of primary focus include contracting for specific targeted clients, reviewing the manner in which resources are allocated to mental health centers, and developing and implementing a set of performance indicators.