PGRMC1 (progesterone receptor membrane component 1) is a highly conserved heme binding protein, which is overexpressed especially in hormone receptor-positive breast cancer and plays an important ...role in breast carcinogenesis. Nevertheless, little is known about the mechanisms by which PGRMC1 drives tumor progression. The aim of our study was to investigate the involvement of PGRMC1 in cholesterol metabolism to detect new mechanisms by which PGRMC1 can increase lipid metabolism and alter cancer-related signaling pathways leading to breast cancer progression.
The effect of PGRMC1 overexpression and silencing on cellular proliferation was examined in vitro and in a xenograft mouse model. Next, we investigated the interaction of PGRMC1 with enzymes involved in the cholesterol synthesis pathway such as CYP51, FDFT1, and SCD1. Further, the impact of PGRMC1 expression on lipid levels and expression of enzymes involved in lipid homeostasis was examined. Additionally, we assessed the role of PGRMC1 in key cancer-related signaling pathways including EGFR/HER2 and ERα signaling.
Overexpression of PGRMC1 resulted in significantly enhanced proliferation. PGRMC1 interacted with key enzymes of the cholesterol synthesis pathway, alters the expression of proteins, and results in increased lipid levels. PGRMC1 also influenced lipid raft formation leading to altered expression of growth receptors in membranes of breast cancer cells. Analysis of activation of proteins revealed facilitated ERα and EGFR activation and downstream signaling dependent on PGRMC1 overexpression in hormone receptor-positive breast cancer cells. Depletion of cholesterol and fatty acids induced by statins reversed this growth benefit.
PGRMC1 may mediate proliferation and progression of breast cancer cells potentially by altering lipid metabolism and by activating key oncogenic signaling pathways, such as ERα expression and activation, as well as EGFR signaling. Our present study underlines the potential of PGRMC1 as a target for anti-cancer therapy.
Decidualization of endometrial cells is the prerequisite for embryo implantation and subsequent placenta formation and is induced by rising progesterone levels following ovulation. One of the hormone ...receptors contributing to endometrial homeostasis is Progesterone Receptor Membrane Component 1 (PGRMC1), a non-classical membrane-bound progesterone receptor with yet unclear function. In this study, we aimed to investigate how PGRMC1 contributes to human decidualization.
We first analyzed PGRMC1 expression profile during a regular menstrual cycle in RNA-sequencing datasets. To further explore the function of PGRMC1 in human decidualization, we implemented an inducible decidualization system, which is achieved by culturing two human endometrial stromal cell lines in decidualization-inducing medium containing medroxyprogesterone acetate and 8-Br-cAMP. In our system, we measured PGRMC1 expression during hormone induction as well as decidualization status upon PGRMC1 knockdown at different time points. We further conferred proximity ligation assay to identify PGRMC1 interaction partners.
In a regular menstrual cycle, PGRMC1 mRNA expression is gradually decreased from the proliferative phase to the secretory phase. In in vitro experiments, we observed that PGRMC1 expression follows a rise-to-decline pattern, in which its expression level initially increased during the first 6 days after induction (PGRMC1 increasing phase) and decreased in the following days (PGRMC1 decreasing phase). Knockdown of PGRMC1 expression before the induction led to a failed decidualization, while its knockdown after induction did not inhibit decidualization, suggesting that the progestin-induced 'PGRMC1 increasing phase' is essential for normal decidualization. Furthermore, we found that the interactions of prohibitin 1 and prohibitin 2 with PGRMC1 were induced upon progestin treatment. Knocking down each of the prohibitins slowed down the decidualization process compared to the control, suggesting that PGRMC1 cooperates with prohibitins to regulate decidualization.
According to our findings, PGRMC1 expression followed a progestin-induced rise-to-decline expression pattern during human endometrial decidualization process; and the correct execution of this expression program was crucial for successful decidualization. Thereby, the results of our in vitro model explained how PGRMC1 dysregulation during decidualization may present a new perspective on infertility-related diseases.
In previous studies, we reported that progesterone receptor membrane component 1 (PGRMC1) is implicated in progestin signaling and possibly associated with increased breast cancer risk upon combined ...hormone replacement therapy. To gain mechanistic insight, we searched for potential PGRMC1 interaction partners upon progestin treatment by co-immunoprecipitation and mass spectrometry. The interactions with the identified partners were further characterized with respect to PGRMC1 phosphorylation status and with emphasis on the crosstalk between PGRMC1 and estrogen receptor α (ERα). We report that PGRMC1 overexpression resulted in increased proliferation of hormone receptor positive breast cancer cell lines upon treatment with a subgroup of progestins including norethisterone and dydrogesterone that promote PGRMC1-phosphorylation on S181. The ERα modulators prohibitin-1 (PHB1) and prohibitin-2 (PHB2) interact with PGRMC1 in dependency on S181-phosphorylation upon treatment with the same progestins. Moreover, increased interaction between PGRMC1 and PHBs correlated with decreased binding of PHBs to ERα and subsequent ERα activation. Inhibition of either PGRMC1 or ERα abolished this effect. In summary, we provide strong evidence that activated PGRMC1 associates with PHBs, competitively removing them from ERα, which then can develop its transcriptional activities on target genes. This study emphasizes the role of PGRMC1 in a key breast cancer signaling pathway which may provide a new avenue to target hormone-dependent breast cancer.
Circulating tumor cells (CTCs) are constantly shed by tumor tissue and can serve as a valuable analyte for a gene expression analysis from a liquid biopsy. However, a high proportion of CTCs can be ...apoptotic leading to rapid mRNA decay and challenging the analysis of their transcriptome. We established a workflow to enrich, to identify, and to isolate single CTCs including the discrimination of apoptotic and non‐apoptotic CTCs for further single CTC transcriptome analysis. Viable tumor cells—we first used cells from breast cancer cell lines followed by CTCs from metastatic breast cancer patients—were enriched with the CellSearch system from diagnostic leukapheresis products, identified by immunofluorescence analysis for neoplastic markers, and isolated by micromanipulation. Then, their cDNA was generated, amplified, and sequenced. In order to exclude early apoptotic tumor cells, staining with Annexin V coupled to a fluorescent dye was used. Annexin V staining intensity was associated with decreased RNA integrity as well as lower numbers of total reads, exon reads, and detected genes in cell line cells and CTCs. A comparative RNA analysis of single cells from MDA‐MB‐231 and MCF7 cell lines revealed the expected differential transcriptome profiles. Enrichment and staining procedures of cell line cells that were spiked into blood had only little effect on the obtained RNA sequencing data compared to processing of naïve cells. Further, the detection of transcripts of housekeeping genes such as GAPDH was associated with a significantly higher quality of expression data from CTCs. This workflow enables the enrichment, detection, and isolation of single CTCs for individual transcriptome analyses. The discrimination of apoptotic and non‐apoptotic cells allows to focus on CTCs with a high RNA integrity to ensure a successful transcriptome analysis.
Alternative splicing and disease Tazi, Jamal; Bakkour, Nadia; Stamm, Stefan
Biochimica et biophysica acta,
01/2009, Letnik:
1792, Številka:
1
Journal Article
Recenzirano
Odprti dostop
Almost all protein-coding genes are spliced and their majority is alternatively spliced. Alternative splicing is a key element in eukaryotic gene expression that increases the coding capacity of the ...human genome and an increasing number of examples illustrates that the selection of wrong splice sites causes human disease. A fine-tuned balance of factors regulates splice site selection. Here, we discuss well-studied examples that show how a disturbance of this balance can cause human disease. The rapidly emerging knowledge of splicing regulation now allows the development of treatment options.
Obesity is a growing worldwide epidemic, and patients classified as obese undergoing gynecologic robotic surgery are at increased risk for surgical complications. This study aimed to evaluate the ...feasibility and outcomes of a surgical safety protocol known as the High BMI Body Mass Index Pathway (HBP) for patients with BMI ≥40 kg/m
undergoing planned robotic hysterectomy. Our primary outcome was the rate of all-cause perioperative complications in patients undergoing surgery with the use of the HBP.
A retrospective cohort study.
An academic teaching hospital.
A total of 138 patients classified as morbidly obese (BMI ≥40 kg/m
) undergoing robotic hysterectomy.
The HBP was developed by a multidisciplinary team and was instituted on January 1, 2016, as a quality improvement project. Patients classified as morbidly obese undergoing robotic hysterectomy after this date were compared with consecutive historical controls.
Seventy-two patients underwent robotic hysterectomies on the HBP and were compared with 66 controls. There were no differences in age, BMI, blood loss, number of comorbidities, or cancer diagnosis. Since the implementation of the HBP, there has been a decrease in anesthesia time (-57.0 minutes; p = .001) and total operating room time (-47.0 min; p = .020), as well as lower estimated blood loss (median 150 mL interquartile range 100-200 vs 200 mL interquartile range 100-300; p = .002) and reduction in overnight hospital admissions (33.3% vs 63.6%; p <.001). In the HBP group, there were fewer all-cause complications (19.4% vs 37.9%; p = .023) and infectious complications (8.3% vs 33.3%; p = .001), and there was no increase in the readmission rates (p = .400). In multivariable analysis, the HBP reduced all-cause complications (odds ratio 0.353; p = .010) after controlling for the covariate (total time in the operating room).
The HBP is a feasible method of optimizing the outcome for patients classified as morbidly obese undergoing major gynecologic surgery. Initiation of the HBP can lead to decreased anesthesia and operating times, all-cause complications, and overnight hospital admissions without increasing readmission rates.
There is a growing family of novel GTPases conserved among higher plants and vertebrates, abbreviated as AIG1, IAP, IMAP, and IAN, respectively. Here, we comparatively analyze the human gene family ...encoding GTPases of the immunity-associated protein family recently re-termed GIMAP. Chromosome 7q36.1 contains, within 300 kb, a
gimap gene cluster with seven functional genes and one pseudogene (h
gimap3). The six genes h
gimap1, h
gimap2, h
gimap4, h
gimap5, h
gimap6, and h
gimap7 encode 33–46 kDa proteins with one GTP-binding domain, whereas h
gimap8 encodes a very unusual 75-kDa protein with three GTP-binding domains. All h
gimap genes except h
gimap2 have orthologs in the mouse. Major expression sites of hgimap mRNAs are the spleen and lymph nodes, but also other organs such as muscle, heart, placenta, and digestive tract display detectable hgimap mRNA levels. The proteins hGIMAP4 and hGIMAP7 can be localized at ER and Golgi apparatus, but not in mitochondria, lysosomes and nuclei. All h
gimap genes were expressed at very low levels—if at all—in diverse cancer cell lines. Our data support the view that the GIMAP proteins are involved in the control of cell survival not only in cells of the immune system as commonly anticipated.
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