Nuclear Progesterone Receptor Research Articles

Estrogen receptor β and epidermal growth factor receptor as early-stage prognostic biomarkers of non-small cell lung cancer

As 20% of stage I NSCLC patients develop recurrent and often incurable cancer, the identification of prognostic markers has a meaningful clinical application. The biological significance of steroid hormone and EGF receptors, able to regulate key physiological functions, remains elusive in NSCLC. Our aim was to investigate the prognostic input of estrogen receptors (ERα, ERβ), progesterone receptors (PR) and EGFR in tumors from 58 stage I NSCLC patients. Antigen expression was analyzed by immunohistochemistry. Prognostic evaluation was performed with the multivariate Cox model. We found that about 70 and 40% of samples expressed ERα or ERβ at cytoplasmic or nuclear level, respectively. Besides, only 12.1% of samples weakly expressed nuclear PR and 62.7% showed membrane EGFR staining. Correlation studies indicated an inverse association between EGFR expression and smoking status (p<0.01). Multivariate studies showed that the lack of nuclear ERβ or the loss of EGFR expression were independent prognosis markers associated with shorter overall survival. We also found that patients whose tumors were negative for these two biomarkers presented the worst outcome. In conclusion, our findings could be useful for selecting stage I NSCLC patients with poor prognosis to apply an earlier treatment that impacts on survival.

Inhibition of the Progesterone Nuclear Receptor during the Bone Linear Growth Phase Increases Peak Bone Mass in Female Mice

Augmentation of the peak bone mass (PBM) may be one of the most effective interventions to reduce the risk of developing osteoporosis later in life; however treatments to augment PBM are currently limited. Our study evaluated whether a greater PBM could be achieved either in the progesterone nuclear receptor knockout mice (PRKO) or by using a nuclear progesterone receptor (nPR) antagonist, RU486 in mice. Compared to their wild type (WT) littermates the female PRKO mice developed significantly higher cancellous and cortical mass in the distal femurs, and this was associated with increased bone formation. The high bone mass phenotype was partially reproduced by administering RU486 in female WT mice from 1–3 months of age. Our results suggest that the inhibition of the nPR during the rapid bone growth period (1–3 months) increases osteogenesis, which results in acquisition of higher bone mass. Our findings suggest a crucial role for progesterone signaling in bone acquisition and inhibition of the nPR as a novel approach to augment bone mass, which may have the potential to reduce the burden of osteoporosis.

Progesterone receptor requires a co-chaperone for signalling in uterine biology and implantation.

Embryo implantation is absolutely dependent on the preparation of the uterus to the receptive stage and attainment by the blastocyst of implantation competency. Co-ordinated effects of progesterone and oestrogen are essential for these processes and determine the window of implantation. In rodents, a generalized stromal edema occurs before blastocyst attachment followed by uterine luminal closure. This leads to apposition of the blastocyst trophectoderm against the luminal epithelium and ultimately attachment. Progesterone is essential for luminal closure, which must occur for successful implantation. More importantly, progesterone is critical for almost every stage of pregnancy, including ovulation, fertilization, implantation, decidualization and pregnancy maintenance. Progesterone exerts its effects on target tissues primarily via nuclear progesterone receptor (PR) whose optimal activity is potentiated by an immunophilin co-chaperone, FK-506 binding protein 4 (FKBP52). While mice lacking PR are infertile due to complete failure of ovulation, fertilization, and implantation, female mice with targeted deletion of the Fkbp52 gene are infertile specifically because of implantation failure resulting from compromised uterine receptivity. This review highlights the evolution of knowledge about progesterone signalling during early pregnancy. Future studies are likely to provide a better understanding of FKBP52-PR signalling in promoting uterine receptivity for implantation and may reveal new targets for improving infertility.

Comparison between steroid binding to membrane progesterone receptor α (mPRα) and to nuclear progesterone receptor: Correlation with physicochemical properties assessed by comparative molecular field analysis and identification of mPRα-specific agonists

Recent results showing that the binding characteristics of 33 steroids for human membrane progesterone receptor alpha (hu-mPRα) differ from those for the nuclear progesterone receptor (nPR) suggest that hu-mPRα-specific agonists can be identified for investigating its physiological functions. The binding affinities of an additional 21 steroids for hu-mPRα were determined to explore the structure–activity relationships in more detail and to identify potent, specific mPRα agonists. Four synthetic progesterone derivatives with methyl or methylene groups on positions 18 or 19, 18a-methylprogesterone (18-CH 3P4, Org OE 64-0), 13-ethenyl-18-norprogesterone (18-CH 2P4, Org 33663-0), 19a-methylprogesterone (19-CH 3P4, Org OD 13-0) and 10-ethenyl-19-norprogesterone (19-CH 2P4, Org OD 02-0), showed similar or higher affinities than progesterone for hu-mPRα and displayed mPRα agonist activities in G-protein and MAP kinase activation assays. All four steroids also bound to the nPR in cytosolic fractions of MCF-7 cells. However, two compounds, 19-CH 2P4 and 19-CH 3P4, showed no nPR agonist activity in a nPR reporter assay and therefore are selective mPRα agonists suitable for physiological investigations. The structure–binding relationships of the combined series of 54 steroids for hu-mPRα deviated strikingly from those of a published set of 60 3-keto or 3-desoxy steroids for nPR. Close correlations were observed between the receptor binding affinities of the steroids and their physicochemical properties calculated by comparative molecular field analysis (CoMFA) for both hu-mPRα and nPR. A comparison of the CoMFA field graphs for the two receptors revealed several differences in the structural features required for binding to hu-mPRα and nPR which could be exploited to develop additional mPR-specific ligands.

Modulation of ATP-induced calcium signaling by progesterone in T47D-Y breast cancer cells

Extracellular ATP activates purinergic (P(2)) receptors with an increase in intracellular calcium and phosphorylation of MAPK. In this study we have investigated the effect of progesterone/progestin on ATP-induced calcium mobilization and phosphorylation of the kinase ERK in the T47D-Y breast cancer cell line that exhibits no detectable nuclear progesterone receptor expression. Brief pretreatment with progesterone/progestin results in a dose dependent inhibition of ATP-induced intracellular calcium mobilization, and inhibition of ERK phosphorylation. Response to a cell impermeable ligand and inhibition of the response by an inactivating antibody suggests a mechanism of action at the plasma membrane. These results in T47D-Y cells strongly suggest that progesterone can act in a rapid non-nuclear manner to inhibit extracellular ATP effects on intracellular calcium mobilization and ERK activation. This research provides an example of progesterone action in a breast cancer cell line lacking expression of the classical nuclear progesterone receptors.

204: Locally produced prostaglandins affect progesterone receptor expression in human pregnancy myometrium

Relaxatory actions of progesterone on the pregnancy myometrium are mediated by the nuclear progesterone receptors, PR-A and PR-B. We previously showed that prostaglandins (PGs) especially PGF2α modulate PR-A and PR-B expression in a human myometrial cell line. The implication of those findings is that locally produced PGs could affect myometrial progesterone responsiveness during pregnancy and in particular induce functional progesterone withdrawal at parturition. The objective of the present study was to determine the effects of locally produced PGs and PGF2α on PR-A and PR-B expression in explant cultures of term human myometrium.

Abstract C23: Therapy of triple-negative breast tumors by targeting the MYC oncogene pathway

Abstract Human breast cancers that lack expression of the estrogen and progesterone nuclear receptors and with low HER2 expression are known as “gtriple-negative” tumors. These are amongst the most aggressive breast cancers and no targeted therapeutics exists for this subclass of human tumors. MYC is a proto-oncogene transcription factor that plays a key role in cell cycle transition of mammalian cells. Increased expression of MYC is estimated to occur in ∼ 20–50% of human breast cancers. Genomic amplification of the MYC locus, 8q24, is correlated with decreased patient survival, and MYC-over-expression in breast cancer cells has been associated with a resistance to anti-hormonal therapies that seek to inhibit ER signaling. Furthermore, MYC has recently been shown to be one of the genes unique to high-grade mammary tumors. Thus, these prior studies suggest that the level of MYC expression might be useful as a biomarker and that the inhibition of MYC function may be a useful therapeutic approach against this difficult-to-treat type of breast cancer. However, while MYC is strongly implicated in breast tumorigenesis, no direct small molecule inhibitors of its transcriptional activity are available in clinical practice. An alternative approach to directly inhibiting MYC is to find “synthetic-lethal” combinations that selectively kill those cells with elevated MYC levels without affecting normal cells. The term “synthetic-lethality” refers to a genetic context in which a mutation in a gene, which itself does not cause lethality, can cause lethality when combined with a mutation in another gene. Using engineered cells, we have previously demonstrated that inhibiting cyclin-dependent kinase 1 (CDK1) can kill cells that over-express MYC. We have now used a panel of breast cancer cell lines established from human patients to show that (1) those breast cancers that are receptor-negative have significantly higher levels of MYC protein, (2) that the MYC pathways are critical for the survival and proliferation of this tumor subset, (3) CDK1 inhibition in triple-negative breast cancer cells results in the initiation of apoptosis mediated by mitochondria-dependent intrinsic pathway, and that (4) this apoptotic event is specifically achieved by altering the balance between pro-survival and pro-apoptotic members of the Bcl-2 family proteins. Our results present a novel treatment strategy for “triple-negative” breast cancers by targeting the MYC pathways via CDK inhibitors. Citation Information: Cancer Res 2009;69(23 Suppl):C23.

Adam12 plays a role during uterine decidualization in mice

In mouse, decidualization is characterized by the proliferation of stromal cells and their differentiation into specialized type of cells (decidual cells) with polyploidy, surrounding the implanting blastocyst. However, the mechanisms involved in these processes remain poorly understood. Using multiple approaches, we have examined the role of Adam12 in decidualization during early pregnancy in mice. Adam12 is spatiotemporally expressed in decidualizing stromal cells in intact pregnant females and in pseudopregnant mice undergoing artificially induced decidualization. In the ovariectomized mouse uterus, the expression of Adam12 is upregulated after progesterone treatment, which is primarily mediated by nuclear progesterone receptor. In a stromal cell culture model, the expression of Adam12 gradually rises with the progression of stromal decidualization, whereas the attenuated expression of Adam12 after siRNA knockdown significantly blocks the progression of decidualization. Our study suggests that Adam12 is involved in promoting uterine decidualization during pregnancy.

Transcriptional effects of progesterone receptor antagonist in rat granulosa cells

Progesterone, acting via the nuclear progesterone receptor (PGR), reduces apoptosis in periovulatory granulosa cells, and is a likely mediator of the anti-atretic actions of LH. The underlying mechanisms, however, have not been clearly defined. In this study, we sought to identify progesterone-mediated transcriptional changes involved in apoptosis regulation. Granulosa cells from immature, gonadotropin-primed female rats were treated in vitro with 100 nM of the PGR antagonist Org 31710. Transcriptional effects were analyzed after 5 and 22 h of incubation using microarrays, and the expression of 85 genes was subsequently measured by quantitative PCR. Follow-up experiments focused on genes related to the functional group "apoptosis". We have identified novel, early gene targets of PGR that may be involved in the control of apoptosis and other biologically significant functions in periovulatory granulosa cells. This study expands our knowledge of events that occur during the processes of ovulation and luteinization.

In Pursuit of Leucine-Rich Repeat-Containing G Protein-Coupled Receptor-5 Regulation and Function in the Uterus

Leucine-rich repeat-containing G protein-coupled receptor (LGR)-5 is a recently identified marker of stem cells in adult intestinal epithelium and hair follicles. Because of this characteristic, we studied the status of Lgr5 expression in the mouse uterus under various conditions. Lgr5 is highly expressed in the uterine epithelium of immature mice and is dramatically down-regulated after the mice resume estrous cycles. Surprisingly, whereas its expression is up-regulated in uteri of ovariectomized mice, the expression is down-regulated by estrogen and progesterone via their cognate nuclear receptors, estrogen receptor-alpha and progesterone receptor, respectively. Using a mouse endometrial cancer model, we also found that Lgr5 is highly expressed in the epithelium during the initial stages of tumorigenesis but is remarkably down-regulated in fully developed tumors. Lgr5 is a downstream target of Wnt signaling in the intestine. Genetic evidence shows that either excessive or absence of Wnt signaling dampens Lgr5 expression in the uterus. Collectively, our results show that Lgr5 expression in the mouse uterine epithelium is unique and dynamically regulated under various physiological and pathological states of the uterus, suggesting that this orphan receptor has important functions in uterine biology. However, identifying definitive uterine function of LGR5 will require further investigation using conditional deletion of uterine Lgr5 because systemic deletion of this gene is neonatally lethal.

Pro- and anticarcinogenic mechanisms of piceatannol are activated dose dependently in MCF-7 breast cancer cells

Estrogenic procarcinogenic effects of piceatannol (PIC) contrast reports about anticarcinogenic activities of PIC. To explain this contradiction, we investigated PIC in estrogen-dependent MCF-7 breast cancer cells and elucidated those cellular mechanisms that correlated with the observed cell effects induced by PIC. Low PIC concentrations (50 nM) induced c-Myc that depended on progesterone receptor (PR) and estrogen receptor (ER). PR-mediated c-Myc induction by PIC was independent of nuclear PR activity but depended on mitogen-activated protein kinase (MAPK) signaling and was associated with an acceleration of cancer cell proliferation. In contrast, 25 μM PIC inhibited deoxynucleotide triphosphate synthesis, activated Chk2 and p38-MAPK and this was accompanied by an attenuation of cancer cell growth. Apoptosis was most probably inhibited due to activation of Akt; however, high PIC concentrations (>100 μM) permitted apoptosis-like cell death in consequence to disruption of orchestrated mitotic signaling. The presented results show for the first time that nanomolar PIC concentrations signal through PR and Erk1/2 and provide a mechanistic explanation why moderate wine consumption-but not other alcoholic beverages-increases the breast cancer risk in women. In contrast, higher PIC concentrations in the micromolar range are considered for adjuvant anticancer therapeutic concepts.

Progesterone stimulates mitochondrial activity with subsequent inhibition of apoptosis in MCF-10A benign breast epithelial cells

The effects of progesterone on breast epithelial cells remain poorly defined with observations showing both proliferative and antiproliferative effects. As an example, progesterone levels correlate with increased epithelial cell proliferation, but there is discordance between the dividing cells and the cells with nuclear progesterone receptor expression. The release of paracrine growth factors from nuclear receptor-positive cells has been postulated as a mechanism, since in vitro studies show a lack of growth effect by progesterone in breast epithelial cells lacking nuclear receptors. This study examined possible nongenomic effects of progesterone in breast epithelia by using MCF-10A cells known to lack nuclear progesterone receptor expression. Treatment for 30-60 min with progesterone or the progestin, R5020, increased mitochondrial activity as shown by an increase in mitochondrial membrane potential (hyperpolarization) with a concordant increase in total cellular ATP. The reaction was inhibited by a specific progesterone receptor antagonist and not affected by the translation inhibitor cycloheximide. Progestin treatment inhibited apoptosis induced by activation of the FasL pathway, as shown by a decrease in sub-G(1) cell fraction during fluorescence-activated cell sorting and a decrease in caspase 3/7 levels. Progestin treatment did not alter the cell cycle over 48 h. Our study demonstrates a nongenomic action of progesterone on benign breast epithelial cells, resulting in enhanced cellular respiration and protection from apoptosis.

Effects of the levonorgestrel-releasing intrauterine system on cell proliferation, Fas expression and steroid receptors in endometriosis lesions and normal endometrium

The objectives of this study were: (i) to evaluate the effects of the levonorgestrel-releasing intrauterine system (LNG-IUS) on both proliferation and apoptosis markers and hormone receptors of the eutopic and ectopic endometrium of women experiencing pain related to endometriosis and (ii) to compare the results with those obtained with GnRH agonist (GnRHa) injections. Pre- and post-treatment endometrium and endometriosis specimens were obtained from 22 women experiencing pain related to endometriosis who were treated with LNG-IUS (n = 11) or GnRHa (n = 11) for 6 months. Changes in the expression of proliferating cell nuclear antigen, Fas, progesterone receptor (PRA) and estrogen receptor alpha (ER-alpha) were analyzed by immunohistochemistry. The cell proliferation index was significantly reduced in the epithelium and stroma of both the eutopic and the ectopic endometrium after treatment with the LNG-IUS and GnRHa. Only LNG-IUS users showed an increased H-score for Fas in the epithelium of the eutopic and ectopic endometrium (P < 0.05). Expression of ER-alpha and PRA by the glandular epithelium was lower in the eutopic endometrium after both treatments, but this reduction was noted in the ectopic endometrium only after LNG-IUS treatments (P < 0.05). No difference was detected between groups for any of the markers. LNG-IUS reduced both cell proliferation and the expression of PRA and ER-alpha and increased Fas expression in the eutopic and ectopic endometrium of patients with endometriosis. Some of these actions were not observed with GnRHa.

Progesterone suppresses the fetal inflammatory response ex vivo

Progesterone supplementation has been shown to be efficacious in preventing preterm birth. We sought to investigate the effects of progesterone on fetal inflammatory responses. Fetal mononuclear cells were isolated from umbilical cord blood and exposed to vehicle or progesterone (P4) for 1 hour prior to lipopolysaccharide (LPS) stimulation. Supernatants were assayed for tumor necrosis factor-alpha. Similar experiments were performed using cyclic adenosine monophosphate (cAMP) and progesterone modulators. The effect of P4 treatment on intracellular cAMP levels was also determined. LPS treatment led to a significant increase in cytokine production by fetal mononuclear cells. Despite the lack of detectable nuclear progesterone receptors, P4 suppressed this inflammatory response. R5020 (progesterone agonist), forskolin (cAMP inducer), and dibutyryl cAMP (cAMP agonist) all achieved immunosuppression. The cAMP antagonist, Rp-cAMP, blocked the inhibitory effect of progesterone. P4 significantly increased intracellular cAMP levels. Progesterone rapidly suppresses the fetal inflammatory response, possibly via nongenomic activation of the cAMP cascade.

Binding of Progesterone to Specific Sites in Isolated Hepatic Cells and Purified Plasma Membrane Fraction

Specific binding for progesterone has been determined in rat hepatocytes and mouse liver purified plasma membranes. The binding is saturable, reversible and temperature dependent. Two types of binding sites have been characterized in hepatocytes. The first is of high affinity and low binding capacity and the other one is of low affinity and high capacity of binding. In plasma membranes one type of specific binding site only exists. These high affinity sites are different from nuclear progesterone receptor, nuclear glucocorticoid receptor, digitalis receptor of Na+, K(+)-ATPase, transcortine and from corticoid binding sites determined previously in plasma membrane. We also have observed that specific progesterone binding to hepatocytes and plasma membrane is independent from the alpha and beta adrenergic receptors and from P-site adenosine receptor.

Paracrine and autocrine functions of the placenta: a key to the success of viviparity?

The evolution of specific nuclear transcriptional regulators has endowed tissues of the reproductive system with responsiveness to small hydrophobic compounds such as steroids. Steroids are widely distributed in Nature and their distribution in prokaryotes and eukaryotes has given rise to the concept that their hormonal role came about by target organ specialization and not by the evolution of steroids themselves. Specific nuclear receptors for progesterone in the uterus are prominent during the establishment and maintenance of pregnancy. Anti-progesterone antagonists which interfere with receptor-mediated DNA activation abrogate pregnancy and thus emphasize the functional importance of the pathways by which the effects of progesterone as an extracellular signal are transduced. Comparative studies show that progesterone itself can be ovarian or placental in origin. This seems to reflect the evolution of different mechanisms of endocrine function rather than any obvious selective advantage being associated with the source of hormone secretion. For this reason, the question of whether the endocrine function of the placenta is obligatory for the adoption of viviparity in mammals is far from certain, and should be considered as an evolutionary option rather than a sine qua non. Of growing importance is the idea that the interaction between trophoblast and endometrial cells controls the degree of invasiveness at implantation and immunoreactivity.(ABSTRACT TRUNCATED AT 250 WORDS)

Expression analysis of steroid hormone receptor mRNAs during zebrafish embryogenesis

We have analyzed by qRT-PCR and/or RT-PCR the abundance and degradation rate of maternal mRNAs for nine steroid hormone receptors and their possible replacement by corresponding embryonic transcripts in both ovulated oocytes and embryos of zebrafish collected at 0, 1, 2, 4, 8, 12, 24 and 48 h post-fertilization (hpf). The mRNAs encoded the nuclear receptors for progesterone ( pr), androgen ( ar), estrogen ( erα, erβ1 and erβ2), glucocorticoids ( gr), mineralocorticoids ( mr) and the membrane progestin receptor-α and β ( mprα and β). gr mRNA was the most abundant maternal transcript in oocytes and early embryos followed by erβ2 and ar mRNAs. They declined during the first 8 hpf, being replaced, thereafter, by the embryonic messengers. erβ1 and mr transcript levels were low until 8 hpf, but increased steadily during embryonic transcription from 24 to 48 hpf. pr transcripts were detectable only in ovulated oocytes and at 24 and 48 hpf. At these stages, there was a slight increase of erα mRNA that initially was very low. mPrα and β mRNAs were expressed in ovulated oocytes and faintly persisted during the first 4 hpf. There was no subsequent embryonic expression of these transcripts. The possible involvement of maternal mRNAs for glucocorticoid and sex hormone receptors in the programming of early zebrafish development is intriguing, since they mainly occur at stages in which gene replication predominates over transcription.

In vitro stimulation of oocyte ovulation in teleosts by gonadotropic and steroid hormones

Published data on in vitro stimulation of oocyte maturation and ovulation by gonadotropic and steroid hormones in different teleost species are reviewed. The involvement of meiosis-inducing steroids, eicosanoids, and nuclear progesterone receptor in the mechanism of ovulation induction is considered.

Progesterone Activates a Progesterone Receptor Membrane Component 1-Dependent Mechanism That Promotes Human Granulosa/Luteal Cell Survival But Not Progesterone Secretion

Objective: We performed this study to test the hypothesis that variation in the lamin a/c gene (LMNA) contributes to milder phenotypes of insulin resistance, hyperandrogenism, and/or metabolic syndrome associated with polycystic ovary syndrome (PCOS). Research Design and Methods: We resequenced the coding region, flanking intronic, and proximal promoter regions of the lamin a/c gene in 43 women with PCOS with evidence of upper-body obesity (waist circumference 88 cm) and identified 56 variants, two of which were nonsynonymous substitutions (lmna11 exon1 E98D; lmna24 exon 7 R455C). We genotyped 53 single-nucleotide polymorphisms (44 identified through resequencing and nine included to maximize informativeness of the entire gene) in 624 index (PCOS) cases and 544 controls of European ancestry. We tested for association between these variants and PCOS. In a subset of individuals, we also tested for association with metabolic syndrome and quantitative traits (body mass index, waist circumference, total testosterone, dehydroepiandrosterone sulfate, fasting glucose and insulin, low-density lipoprotein, and TTG). Results: After correction for multiple testing, none of the variants showed significant evidence for association with PCOS, the metabolic syndrome, or any of the quantitative traits tested. Conclusions: Whereas these studies cannot exclude the role of genetic variation in the lamin a/c gene in isolated cases of PCOS, we can conclude that common variation in the lamin a/c gene does not contribute to the etiology of PCOS in women of European ancestry. Progesterone Activates a Progesterone Receptor Membrane Component 1-Dependent Mechanism That Promotes Human Granulosa/Luteal Cell Survival But Not Progesterone Secretion John J. Peluso, Xiufang Liu, Anna Gawkowska, and Erika Johnston-MacAnanny (J Clin Endocrinol Metab, published May 5, 2009, 10.1210/jc.2009-0147) ABSTRACT Context: Progesterone (P4) promotes its own secretion and the survival of human granulosa/luteal (GL) cells.Context: Progesterone (P4) promotes its own secretion and the survival of human granulosa/luteal (GL) cells. Objective: The objective of these studies was to determine whether progesterone receptor membrane component-1 (PGRMC1) mediates P4’s actions. Design: In vitro studies were conducted on GL cells from women undergoing in vitro fertilization and GL5 cells, which are derived from GL cells. Setting and Patients: GL cells were obtained from women undergoing fertility treatment at a university-based clinic and used for in vitro studies. Main Outcome Measures: PCR, Western blot, and immunocytochemistry were used to detect various progestin binding proteins. H-P4 binding kinetics were assessed on partially purified PGRMC1. Apoptosis was determined after culture by either TUNEL or DAPI staining. P4 was measured by an ELISA assay. PGRMC1 was depleted using small interfering RNA. Results: GL and GL5 cells expressed several P4 binding proteins including the nuclear progesterone receptor (PGR), progestin/ adipoQ receptors (PAQR 7, 8, and 5) and PGRMC1. Ligand binding studies revealed that both P4 and the progestin, R5020, bound PGRMC1 with an EC50 of approximately 10 nM. Interestingly, P4 inhibited apoptosis at concentrations in the 10 nM range, whereas R5020 stimulated P4 secretion at concentrations of at lease 16 M. Depleting PGRMC1 attenuated P4’s antiapoptotic action but failed to influence R5020-induced P4 secretion. T R A N S L A T I O N A L H I G H L I G H T S F R O M J C E M

Evidence for the existence of an oligomeric, non-DNA-binding complex of the progesterone receptor in the cytoplasm.

Steroid receptors are found as a hetero-oligomeric complex in cell extracts. Due to the dynamic interaction between receptor-associated proteins and receptors, it is difficult to study the oligomeric complex in living cells. Here this was attempted in cells in which the interaction was stabilized by introducing molybdate into the cells or by incubating the cells at low temperature. The complex was studied with an antibody (aD) recognizing only the dissociated form of the chicken progesterone receptor (PR) and with antibodies (PR22, PR6). Recognizing also oligomeric forms of the receptor. When wild-type chicken PR was transfected, all antibodies showed nuclear staining. Molybdate or cold treatment of cells resulted in cytoplasmic accumulation of the PR as detected with PR22/PR6. aD, however, stained predominantly the nuclear PR in treated cells. These findings suggest that when the oligomeric complex of the PR is stabilized in intact cells in vivo and then crosslinked with paraformaldehyde, a portion of the cytoplasmic receptor is seen as an oligomeric complex, whereas, in the nucleus, most, if not all receptor molecules are in dissociated form.