Differential regulation of SGK1 by progesterone activates AP-1/NDRG1 genomic axis in PR-positive and negative breast cancer cells
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: E-MTAB-6742 | BioStudies |
REPOSITORIES: biostudies
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Project description:Pre-operative progesterone intervention has been shown to confer a survival benefit to breast cancer patients independent of their progesterone receptor (PR) status, raising a question about how progesterone affects the outcome of PR-negative cells. Here, we identify up-regulation of a Serum- and glucocorticoid-regulated kinase gene, SGK1 and an N-Myc Downstream Regulated Gene 1, NDRG1, along with down-regulation of miR-29a and miR-101-1 targeting 3’UTR region of SGK1, to differential extents in a PR dependent manner in breast cancer cells. We further demonstrate a novel dual-phase transcriptional and post-transcriptional regulation of SGK1 in response to progesterone leading to up-regulation of a tumor metastasis suppressor gene, NDRG1, mediated by a set of AP-1 network genes. The NDRG1 further inactivates a set of kinases impeding the invasion and migration of breast cancer cells. In summary, we propose a model for the mode of action of progesterone in breast cancer deciphering the molecular basis of a randomized clinical trial studying the effect of progesterone in breast cancer with a potential to improve the prognosis of breast cancer patients for receiving pre-operative progesterone treatment.
2018-10-19 | E-MTAB-6742 | ArrayExpress
Project description:Preoperative progesterone intervention has been shown to confer a survival benefit to breast cancer patients independently of their progesterone receptor (PR) status. This observation raises the question how progesterone affects the outcome of PR-negative cancer. Here, using microarray and RNA-Seq-based gene expression profiling and ChIP-Seq analyses of breast cancer cells, we observed that the serum- and glucocorticoid-regulated kinase gene (<i>SGK1</i>) and the tumor metastasis-suppressor gene N-Myc downstream regulated gene 1 (<i>NDRG1</i>) are up-regulated and that the microRNAs <i>miR-29a</i> and <i>miR-101-1</i> targeting the 3'-UTR of <i>SGK1</i> are down-regulated in response to progesterone. We further demonstrate a dual-phase transcriptional and post-transcriptional regulation of <i>SGK1</i> in response to progesterone, leading to an up-regulation of <i>NDRG1</i> that is mediated by a set of genes regulated by the transcription factor AP-1. We found that <i>NDRG1</i>, in turn, inactivates a set of kinases, impeding the invasion and migration of breast cancer cells. In summary, we propose a model for the mode of action of progesterone in breast cancer. This model helps decipher the molecular basis of observations in a randomized clinical trial of the effect of progesterone on breast cancer and has therefore the potential to improve the prognosis of breast cancer patients receiving preoperative progesterone treatment.
| S-EPMC6298595 | BioStudies
Project description:The protein N-myc down-regulated gene 1 (NDRG1) represses tumour metastasis. It is phosphorylated at several sites by serum and glucocorticoid-regulated kinase 1 (SGK1). Here we show that NDRG1 is also regulated by the oncogenic MAP kinase-interacting kinase (MNK) pathway, a target for cancer therapy.Inhibiting MNKs increases the expression of NDRG1 protein and mRNA in breast cancer cells. MNK inhibition also decreases the phosphorylation of NDRG1. Phosphorylation of NDRG1 is reduced in cells lacking MNK1, but not MNK2-knockout cells, indicating that NDRG1 phosphorylation is a specific target for MNK1. However, MNK1 cannot directly phosphorylate NDRG1 in vitro, indicating that additional signalling connections are involved. Taken together, our data indicate that MNK signaling regulates NDRG1 at transcriptional and post-translational levels.We show that SGK1 phosphorylates MNK1 at a conserved site, which represses its activity. NDRG1, SGK1 and the MNKs are implicated in cell migration and metastasis. As expected, knocking-down NDRG1 promoted cell migration. However, whereas MNK inhibition impairs these processes irrespective of NDRG1 levels, SGK inhibition only did so in NDRG1-depleted cells. Thus, MNKs and SGK affect migration/invasion through distinct mechanisms.Our data reveal several novel connections between signalling pathways important for tumour biology.
| S-EPMC5542254 | BioStudies
Project description:Adipocytes play a central role in maintaining metabolic homeostasis in the body. Differentiation of adipocyte precursor cells requires the transcriptional activity of peroxisome proliferator-activated receptor-? (Ppar?) and CCAAT/enhancer binding proteins (C/Ebps). Transcriptional activity is regulated by signaling modules activated by a plethora of hormones and nutrients. Mechanistic target of rapamacin complexes (mTORC) 1 and 2 are central for the coordination of hormonal and nutritional inputs in cells and are essential for adipogenesis. Serum glucocorticoid kinase 1 (Sgk1)-dependent phosphorylation of N-Myc downstream-regulated gene 1 (Ndrg1) is a hallmark of mTORC2 activation in cells. Moreover, Ppar? activation promotes Ndrg1 expression. However, the impact of Ndrg1 on adipocyte differentiation and function has not yet been defined. Here, we show that Ndrg1 expression and its Sgk1-dependent phosphorylation are induced during adipogenesis. Consistently, we demonstrate that Ndrg1 promotes adipocyte differentiation and function by inducing Ppar? expression. Additionally, our results indicate that Ndrg1 is required for C/Ebp? phosphorylation. Moreover, we found that Ndrg1 phosphorylation by Sgk1 promotes adipocyte formation. Taken together, we show that induction of Ndrg1 expression by Ppar? and its phosphorylation by Sgk1 kinase are required for the acquisition of adipocyte characteristics by precursor cells.
| S-EPMC5543145 | BioStudies
Project description:Glutamate receptors play an important role in the function of astrocytes. Among their tasks is the regulation of gliotransmission, gene expression and exocytosis of the tissue-type plasminogen activator (tPA), which has an enhancing effect on N-methyl-D-aspartate (NMDA) receptors and thus prevent over-excitation of neighboring neurons. The kainate receptor GluK2, which is expressed in neurons and astrocytes, is under tight regulation of the PI3-kinase SGK pathway as shown in neurons. SGK1 targets include N-myc downstream-regulated genes (NDRGs) 1 and 2 (NDRG1, NDRG2), proteins with elusive function. In the present study, we analyzed the effects of SGK1, NDRG1, and NDRG2 on GluK2 current amplitude and plasma membrane localization in astrocytes and heterologous expression. We demonstrate that NDRG1 and NDRG2 themselves have no effect on GluK2 current amplitudes in heterologous expressed ion channels. However, when NDRG2 is coexpressed with GluK2 and SGK1, the stimulating effect of SGK1 on GluK2 is suppressed both in heterologous expression and in astrocytes. Here, we reveal a new negative feedback mechanism, whereby GluK2 stimulation by SGK1 is regulated by parallel phosphorylation of NDRG2. This regulation of GluK2 by SGK1 and NDRG2 in astrocytes may play an important role in gliotransmission, modulation of gene expression and regulation of exocytosis of tPA.
| S-EPMC4594022 | BioStudies
Project description:The majority of human cancers harbour mutations promoting activation of the Akt protein kinase, and Akt inhibitors are being evaluated in clinical trials. An important question concerns the understanding of the innate mechanisms that confer resistance of tumour cells to Akt inhibitors. SGK (serum- and glucocorticoid-regulated kinase) is closely related to Akt and controlled by identical upstream regulators {PI3K (phosphoinositide 3-kinase), PDK1 (phosphoinositide-dependent kinase 1) and mTORC2 [mTOR (mammalian target of rapamycin) complex 2]}. Mutations that trigger activation of Akt would also stimulate SGK. Moreover, Akt and SGK possess analogous substrate specificities and are likely to phosphorylate overlapping substrates to promote proliferation. To investigate whether cancers possessing high SGK activity could possess innate resistance to Akt-specific inhibitors (that do not target SGK), we analysed SGK levels and sensitivity of a panel of breast cancer cells towards two distinct Akt inhibitors currently in clinical trials (AZD5363 and MK-2206). This revealed a number of Akt-inhibitor-resistant lines displaying markedly elevated SGK1 that also exhibited significant phosphorylation of the SGK1 substrate NDRG1 [N-Myc (neuroblastoma-derived Myc) downstream-regulated gene 1]. In contrast, most Akt-inhibitor-sensitive cell lines displayed low/undetectable levels of SGK1. Intriguingly, despite low SGK1 levels, several Akt-inhibitor-sensitive cells showed marked NDRG1 phosphorylation that was, unlike in the resistant cells, suppressed by Akt inhibitors. SGK1 knockdown markedly reduced proliferation of Akt-inhibitor-resistant, but not -sensitive, cells. Furthermore, treatment of Akt-inhibitor-resistant cells with an mTOR inhibitor suppressed proliferation and led to inhibition of SGK1. The results of the present study suggest that monitoring SGK1 levels as well as responses of NDRG1 phosphorylation to Akt inhibitor administration could have a use in predicting the sensitivity of tumours to compounds that target Akt. Our findings highlight the therapeutic potential that SGK inhibitors or dual Akt/SGK inhibitors might have for treatment of cancers displaying elevated SGK activity.
| S-EPMC3671793 | BioStudies
Project description:Pre-operative progesterone treatment of breast cancer has been shown to confer survival benefits to patients independent of their progesterone receptor (PR) status. The underlying mechanism and the question whether such an effect can also be observed in PR negative breast cancer cells remain to be resolved.We performed proteome profiling of PR-positive and PR-negative breast cancer cells in response to progesterone using a phospho-kinase array platform. Western blotting was used to validate the results. Cell-based phenotypic assays were conducted using PR-positive and PR-negative breast cancer cells to assess the effect of progesterone.We found that progesterone induces de-phosphorylation of 12 out of 43 kinases tested, which are mostly involved in cellular invasion and migration regulation. Consistent with this observation, we found through cell-based phenotypic assays that progesterone inhibits the invasion and migration of breast cancer cells independent of their PR status.Our results indicate that progesterone can inhibit breast cancer cell invasion and migration mediated by the de-phosphorylation of kinases. This inhibition appears to be independent of the PR status of the breast cancer cells. In a broader context, our study may provide a basis for an association between progesterone treatment and recurrence reduction in breast cancer patients, thereby providing a lead for modelling a randomized in vitro study.
2017-01-01 | S-EPMC5537311 | BioStudies
Project description:BACKGROUND: Cancer cell differentiation is an important characteristic of malignant tumor and has a great impact on prognosis and therapeutic decision for patients. The N-myc downstream regulated gene 1 (NDRG1), a putative tumor suppression gene, is involved in the regulation of human cell differentiation and metastasis in various cancers. Changes in the status of methylation of the NDRG1 gene have not been studied in detail in human breast cancer. RESULTS: The MDA-MB-231 breast tumor cell line could express NDRG1. However, it was only expressed after treatment with 5-Aza-2'-deoxycytidine (AZA) in T47D cell line, which revealed that NDRG1 expression could modulated by DNA methylation. Therefore, the fragment surrounding the transcript start site of NDRG1 gene promoter was cloned after sodium bisulfite DNA treatment. A high density (66%) of methylation for human NDRG1 gene promoter was detected in T47D; however, there was only 16% of methylated CpG dinucleotides in the NDRG1 gene promoter in MDA-MB-231. DNA methylation in the NDRG1 promoter was detected in 31.1% of primary breast cancer samples. Furthermore, the NDRG1 promoter methylation correlated with the Tumor Node Metastasis (TNM) at stage III/IV, metastasis, lymph invasion, moderate and poor histological grade in the breast cancer patients. CONCLUSION: These findings suggest that the DNA methylation status of NDRG1 gene may play an important role in the pathogenesis and/or development of breast cancer, and the expression could be regulated by aberrant DNA methylation.
| S-EPMC3751627 | BioStudies
Project description:BACKGROUND:Altered lipid metabolism is an emerging hallmark of aggressive breast cancers. The N-myc downstream regulated gene (NDRG1) gene plays a critical role in peripheral nervous system myelination, as inactivating mutations cause severe demyelinating neuropathy. In breast cancer, elevated NDRG1 expression has been linked to clinical outcomes, but its functional role in breast cancer physiology has remained unclear. METHODS:A meta-analysis of NDRG1 expression in multiple large publicly available genomic databases was conducted. Genome-wide expression correlation and Cox proportional hazards and Kaplan-Meier modeling of clinical outcomes associated with elevated expression were assessed. To study NDRG1 function, gene silencing and overexpression phenotypic studies were carried out in a panel of cell lines representing all major breast cancer molecular subtypes. Changes in cell proliferation, morphology, and neutral lipid accumulation due to altered NDRG1 expression were assessed by high throughput, quantitative microscopy. Comprehensive lipidomics mass spectrometry was applied to characterize global changes in lipid species due to NDRG1 silencing. Labeled fatty acids were used to monitor cellular fatty acid uptake and subcellular distribution under nutrient replete and starvation culture conditions. RESULTS:NDRG1 overexpression correlated with glycolytic and hypoxia-associated gene expression, and was associated with elevated rates of metastasis and patient mortality. Silencing NDRG1 reduced cell proliferation rates, causing lipid metabolism dysfunction including increased fatty acid incorporation into neutral lipids and lipid droplets. Conversely, NDRG1 expression minimized lipid droplet formation under nutrient replete and starvation conditions. CONCLUSIONS:Here we report that NDRG1 contributes to breast cancer aggressiveness by regulating the fate of lipids in cells that exhibit an altered lipid metabolic phenotype. In line with its role in promoting myelination and its association with altered metabolism in cancer, our findings show that NDRG1 is a critical regulator of lipid fate in breast cancer cells. The association between NDRG1 and poor prognosis in breast cancer suggests it should play a more prominent role in patient risk assessment. The function of NDRG1 in breast cancer lipid metabolism may represent a promising therapeutic approach in the future.
| S-EPMC6001025 | BioStudies
Project description:Hypoxia can lead to solid tumor aggressiveness by driving multiple signaling pathways. Long non-coding RNAs respond to several extrinsic stimuli, causing changes in cancer cells by participating in multiple steps of gene expression. However, genomic profiling of long non-coding RNAs regulated by oxygen in breast cancer remained unclear. Therefore, the aims of this study were to identify oxygen-responsive long non-coding RNAs in breast cancer cells, and to delineate their regulatory mechanisms. The expression profiling of long non-coding RNAs in breast cancer cells growing under normoxic, hypoxic, and re-oxygenated conditions was examined using next-generation sequencing technology. Four hundred and seventy-two lncRNAs oxygen-responsive lncRNAs were identified. After examining the top three differentially expressed lncRNAs in hypoxia, we selected N-Myc Downstream Regulated Gene 1-Overlapping 1 (NDRG1-OT1) for further study, especially the most responsive isoform, NDRG1-OT1_v4. We overexpressed NDRG1-OT1_v4 under normoxia and performed microarray analysis to identify 108 NDRG1-OT1_v4 regulated genes and their functions. Among these genes, we found that both NDRG1 mRNA expression and NDRG1 protein levels were inhibited by NDRG1-OT1_v4. Finally, we used co-immunoprecipitation to show that NDRG1-OT1_v4 destabilizes NDRG1 by promoting ubiquitin-mediated proteolysis. Our findings reveal a new type of epigenetic regulation of NDRG1 by NDRG1-OT1_v4 in breast cancer cells.
| S-EPMC5828211 | BioStudies