CEBPB

Gene Summary

Gene:CEBPB; CCAAT/enhancer binding protein (C/EBP), beta
Aliases: TCF5, IL6DBP, NF-IL6, C/EBP-beta
Location:20q13.1
Summary:This intronless gene encodes a transcription factor that contains a basic leucine zipper (bZIP) domain. The encoded protein functions as a homodimer but can also form heterodimers with CCAAT/enhancer-binding proteins alpha, delta, and gamma. Activity of this protein is important in the regulation of genes involved in immune and inflammatory responses, among other processes. The use of alternative in-frame AUG start codons results in multiple protein isoforms, each with distinct biological functions. [provided by RefSeq, Oct 2013]
Databases:OMIM, VEGA, HGNC, Ensembl, GeneCard, Gene
Protein:CCAAT/enhancer-binding protein beta
HPRD
Source:NCBIAccessed: 20 August, 2015

Ontology:

What does this gene/protein do?
Show (31)
Pathways:What pathways are this gene/protein implicaed in?
Show (1)

Cancer Overview

Research Indicators

Publications Per Year (1990-2015)
Graph generated 20 August 2015 using data from PubMed using criteria.

Literature Analysis

Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic.

  • Squamous Cell Carcinoma
  • Messenger RNA
  • ras Proteins
  • Bladder Cancer
  • Translocation
  • Cervical Cancer
  • siRNA
  • Tumor Markers
  • Cell Proliferation
  • Gene Expression
  • Base Sequence
  • Cell Differentiation
  • Neoplasm Proteins
  • Oligonucleotide Array Sequence Analysis
  • Oncogene Fusion Proteins
  • Proto-Oncogene Proteins
  • Promoter Regions
  • Cancer Gene Expression Regulation
  • Chromosome 20
  • Extracellular Signal-Regulated MAP Kinases
  • Cell Line
  • Polymerase Chain Reaction
  • fms-Like Tyrosine Kinase 3
  • CCAAT-Enhancer-Binding Proteins
  • Gene Expression Profiling
  • Breast Cancer
  • MicroRNAs
  • Acute Myeloid Leukaemia
  • V(D)J Recombination
  • p53 Protein
  • Molecular Sequence Data
  • Interleukin-1beta
  • RNA Interference
  • CCAAT-Enhancer-Binding Protein-beta
  • Vascular Endothelial Growth Factor Receptor-2
  • Phosphorylation
  • Transcription Factors
  • MAP Kinase Signaling System
  • Neoplastic Cell Transformation
  • NF-kappa B
Tag cloud generated 20 August, 2015 using data from PubMed, MeSH and CancerIndex

Specific Cancers (4)

Data table showing topics related to specific cancers and associated disorders. Scope includes mutations and abnormal protein expression.

Note: list is not exhaustive. Number of papers are based on searches of PubMed (click on topic title for arbitrary criteria used).

Latest Publications: CEBPB (cancer-related)

Baron VT, Pio R, Jia Z, Mercola D
Early Growth Response 3 regulates genes of inflammation and directly activates IL6 and IL8 expression in prostate cancer.
Br J Cancer. 2015; 112(4):755-64 [PubMed] Article available free on PMC after 17/02/2016 Related Publications
BACKGROUND: Transcription factor EGR3 (Early Growth Response 3) is a little-studied member of the EGR family that is highly expressed in human prostate tumours compared with normal tissue. Its function in prostate cancer, however, is unknown.
METHODS: Stable shRNA silencing was achieved in naturally overexpressing prostate cancer cells, followed by Affymetrix expression analysis. Fold changes of ⩾2 and ⩽-2 were considered valid and t-tests P-values of ⩽0.01 were considered statistically significant. Potential EGR3 target genes were validated by real-time qPCR, chromatin immunoprecipitation, and gain-of-function experiments. Promoter analysis confirmed the presence of consensus binding sites in the promoters of target genes.
RESULTS: Early Growth Response 3 regulates the expression of ∼330 genes, 35% of which are involved in immune responses and inflammatory processes, and 15% crosstalk with the NF-κB signalling pathway. In particular, EGR3 induces the expression of over 50 secreted cytokines, growth factors, and matrix remodelling factors. Two interleukins of great relevance to prostate cancer, IL6 and IL8, were further validated as EGR3 target genes: both promoters contain EGR consensus binding sites and are pulled down in intact cells by EGR3 chromatin immunoprecipitation. Silencing of EGR3 decreased IL6 and IL8 expression, whereas overexpression of EGR3 in nontransformed cells induced IL6 and IL8 expression.
CONCLUSIONS: Chronic inflammation plays a critical role in prostate cancer and elevated production of pro-inflammatory cytokines IL8 and IL6, in particular, contributes to disease progression and to the onset of castration resistance. It is shown for the first time that EGR3 is involved in the upregulation of both IL6 and IL8. Together with our previous observation that EGR3 is highly expressed in prostate tumours compared with normal tissue and strongly correlates with IL6 and IL8 expression in clinical samples, the present study suggests that EGR3 promotes excessive production of IL6 and IL8 observed during the progression of prostate cancer.

Liu F, Ji F, Ji Y, et al.
Dissecting the mechanism of colorectal tumorigenesis based on RNA-sequencing data.
Exp Mol Pathol. 2015; 98(2):246-53 [PubMed] Related Publications
OBJECTIVE: This study aimed to identify the differentially expressed genes (DEGs), mutated genes and fusion genes in colorectal cancer.
MATERIALS AND METHODS: RNA-sequencing data (ID: SRP009386) from cancerous, paracancerous non-tumor and distant normal tissue from one Chinese patient with stage III colorectal cancer were downloaded from Sequence Read Archive. Quality control was checked using FastQC, followed by sequence alignment against the hg19 reference genome using TopHat v1.3.3. The expression levels were quantified using Cufflinks, followed by DEGs screening using NOISeq. Enrichment analysis was performed using DAVID. Transcription factors were screened using TRANSFA. Mutated loci were identified using SAMTools and VCFTools. Gene fusion events were detected by TopHat-fusion.
RESULTS: In total 2440, 1887 and 834 DEGs were respectively detected in cancerous vs. normal tissue, cancerous vs. paracancerous tissue and paracancerous vs. normal tissue. The up-regulated genes from cancerous and paracancerous tissue compared with normal tissue were enriched in "extracellular matrix receptor interaction" and "focal adhesion pathway" as well as some biological processes except for "negative regulation of programmed cell death" uniquely presenting in cancer. Dysregulated transcription factors including SOX4, BCL6, CEBPB and MSX2 were enriched in the unique biological process. Trp53 was identified with one mutated locus 7577142 (C → T) on chromosome 17. BCL6 also experienced missense mutation. Additionally, COL1A1-PPP2R2C and EXPH5-COL1A2 were observed fusion genes in cancer tissue.
CONCLUSIONS: The unique biological process in cancer tissue may be the cause for colorectal carcinogenesis. The screened transcription factors, mutated genes and fusion genes may contribute to the progression of colorectal cancer.

Boukhari A, Alhosin M, Bronner C, et al.
CD47 activation-induced UHRF1 over-expression is associated with silencing of tumor suppressor gene p16INK4A in glioblastoma cells.
Anticancer Res. 2015; 35(1):149-57 [PubMed] Related Publications
CD47, an integrin-associated protein is over-expressed in several tumors including glioblastomas. Activation of CD47 induces proliferation of human astrocytoma cells but not normal astrocytes via an Akt-dependent way. However, the pathways mediating this process are still unknown. The epigenetic integrator UHRF1 (Ubiquitin-like containing PHD and RING Finger 1) is over-expressed in various cancers and plays a vital role in the silencing of numerous tumor suppressor genes including p16(INK4A), thereby promoting cell proliferation. The aim of the present study was to investigate the role of UHRF1 and p16(INK4A) in CD47-induced effects. Herein we showed that activation of CD47 in human astrocytoma cell lines U87 and CCF- STTG1 (Grade IV), up-regulated the expression of UHRF1 with subsequent down-regulation of p16(INK4A), thus promoting cell proliferation. Blockage of CD47 using a blocking antibody down-regulated UHRF1 expression, accompanied by a re-expression of p16(INK4A), conducting to decreased cell proliferation in both cancer cell lines. Neither CD47 activation nor its blocking has any effect on UHRF1/p16(INK4A) expression in normal human astrocytes. Depletion of CD47 in the U87 cell line resulted in down-regulation of UHRF1. We also found that CD47 activated the inflammatory genes IL-6, IL-7 and MCP-1 by a NF-κB-dependent mechanism in human astrocytoma but not in normal astrocytes. In conclusion, the present findings indicate that CD47 activation increases expression of UHRF1 and suggest, for the first time, that CD47 regulates the epigenetic code by targeting UHRF1. This could represent a new pathway towards cell proliferation and metastasis.

Salotti J, Sakchaisri K, Tourtellotte WG, Johnson PF
An Arf-Egr-C/EBPβ pathway linked to ras-induced senescence and cancer.
Mol Cell Biol. 2015; 35(5):866-83 [PubMed] Article available free on PMC after 01/09/2015 Related Publications
Oncogene-induced senescence (OIS) protects normal cells from transformation by Ras, whereas cells lacking p14/p19(Arf) or other tumor suppressors can be transformed. The transcription factor C/EBPβ is required for OIS in primary fibroblasts but is downregulated by H-Ras(V12) in immortalized NIH 3T3 cells through a mechanism involving p19(Arf) loss. Here, we report that members of the serum-induced early growth response (Egr) protein family are also downregulated in 3T3(Ras) cells and directly and redundantly control Cebpb gene transcription. Egr1, Egr2, and Egr3 recognize three sites in the Cebpb promoter and associate transiently with this region after serum stimulation, coincident with Cebpb induction. Codepletion of all three Egrs prevented Cebpb expression, and serum induction of Egrs was significantly blunted in 3T3(Ras) cells. Egr2 and Egr3 levels were also reduced in Ras(V12)-expressing p19(Arf) null mouse embryonic fibroblasts (MEFs), and overall Egr DNA-binding activity was suppressed in Arf-deficient but not wild-type (WT) MEFs, leading to Cebpb downregulation. Analysis of human cancers revealed a strong correlation between EGR levels and CEBPB expression, regardless of whether CEBPB was increased or decreased in tumors. Moreover, overexpression of Egrs in tumor cell lines induced CEBPB and inhibited proliferation. Thus, our findings identify the Arf-Egr-C/EBPβ axis as an important determinant of cellular responses (senescence or transformation) to oncogenic Ras signaling.

Zheng R, Wang X, Studzinski GP
1,25-Dihydroxyvitamin D3 induces monocytic differentiation of human myeloid leukemia cells by regulating C/EBPβ expression through MEF2C.
J Steroid Biochem Mol Biol. 2015; 148:132-7 [PubMed] Article available free on PMC after 01/04/2016 Related Publications
Myogenic enhancer factor2 (Mef2) consists of a family of transcription factors involved in morphogenesis of skeletal, cardiac and smooth muscle cells. Among the four isoforms (Mef2A, 2B, 2C, and 2D), Mef2C was also found to play important roles in hematopoiesis. At myeloid progenitor level, Mef2C expression favors monocytic differentiation. Previous studies from our laboratory demonstrated that ERK5 was activated in 1,25-dihydroxyvitamin D3 (1,25D)-induced monocytic differentiation in AML cells and ERK5 activation was accompanied by increased Mef2C phosphorylation. We therefore examined the role of Mef2C in 1,25D-induced monocytic differentiation in AML cell lines (HL60, U937 and THP1) and found that knockdown of Mef2C with small interfering RNA (siRNA) significantly decreases the expression of the monocytic marker, CD14, without affecting the expression of the general myeloid marker, CD11b. CCAAT/enhancer-binding protein (C/EBP) β, which can bind to CD14 promoter and increase its transcription, has been shown to be the downstream effector of 1,25D-induced monocytic differentiation in AML cells. When Mef2C was knocked down, expression of C/EBPβ was reduced at both mRNA and protein levels. The protein expression levels of cell cycle regulators, p27(Kip1) and cyclin D1, were not affected by Mef2C knockdown, nor the monopoiesis related transcription factor, ATF2 (activating transcription factor 2). Thus, we conclude that 1,25D-induced monocytic differentiation, and CD14 expression in particular, are mediated through activation of ERK5-Mef2C-C/EBPβ signaling pathway, and that Mef2C does not seem to modulate cell cycle progression.

Anand S, Ebner J, Warren CB, et al.
C/EBP transcription factors in human squamous cell carcinoma: selective changes in expression of isoforms correlate with the neoplastic state.
PLoS One. 2014; 9(11):e112073 [PubMed] Article available free on PMC after 01/04/2016 Related Publications
The CCAAT/Enhancer Binding Proteins (C/EBPs) are a family of leucine-zipper transcription factors that regulate physiological processes such as energy metabolism, inflammation, cell cycle, and the development and differentiation of several tissues including skin. Recently, a role for C/EBPs in tumor cell proliferation and differentiation has been proposed, but the incomplete characterization in the literature of multiple translational isoforms of these proteins has made interpretation of these roles difficult. Therefore, we have carefully reexamined C/EBP isoform expression in human non-melanoma skin cancers. C/EBPα, C/EBPβ, and C/EBPδ were analyzed histologically in squamous cell carcinomas (SCC). The individual isoforms of C/EBPα and C/EBPβ were examined by immunofluorescent digital imaging, western blotting and DNA binding activity (electrophoretic mobility shift analysis). Expression of all C/EBP family proteins was decreased in SCC tumors. Suppression was greatest for C/EBPα, less for C/EBPβ, and least for C/EBPδ. Western analyses confirmed that C/EBPα p42 and p30 isoforms were decreased. For C/EBPβ, only the abundant full-length isoform (C/EBPβ-1, LAP*, 55 kD) was reduced, whereas the smaller isoforms, C/EBPβ-2 (LAP, 48 kD) and C/EBPβ-3 (LIP, 20 kD), which are predominantly nuclear, were significantly increased in well- and moderately-differentiated SCC (up to 14-fold for C/EBPβ-3). These elevations correlated with increases in PCNA, a marker of proliferation. Although C/EBPβ displayed increased post-translational modifications in SCC, phosphorylation of C/EBPβ-1 (Thr 235) was not altered. C/EBP-specific DNA binding activity in nuclear and whole-cell extracts of cultured cells and tumors was predominantly attributable to C/EBPβ. In summary, two short C/EBPβ isoforms, C/EBPβ-2 and C/EBPβ-3, represent strong candidate markers for epithelial skin malignancy, due to their preferential expression in carcinoma versus normal skin, and their strong correlation with tumor proliferation.

Liu X, Liu K, Qin J, et al.
C/EBPβ promotes angiogenesis through secretion of IL-6, which is inhibited by genistein, in EGFRvIII-positive glioblastoma.
Int J Cancer. 2015; 136(11):2524-34 [PubMed] Related Publications
To study the mechanisms underlying the IL-6-promoted angiogenic microenvironment in EGFRvIII-positive glioblastoma, VEGF expression in EGFRvIII-positive/negative tumors was determined by optical molecular imaging. Next, the HUVEC tube formation assay, Western blot, qPCR, RNA silencing, chromatin immunoprecipitation, luciferase reporter and ELISA assays were performed to examine the role of IL-6 and C/EBPβ in the formation of the angiogenic microenvironment in EGFRvIII-positive tumors. Finally, in vitro and in vivo genistein treatment experiments were conducted to challenge the interaction between the IL-6 promoter and C/EBPβ. Optical imaging revealed greater VEGF expression in EGFRvIII-positive tumor-bearing mice, suggesting an angiogenic microenvironment. In vitro experiments demonstrated that C/EBPβ-mediated regulation of IL-6 was indispensable for maintenance of this angiogenic microenvironment. In contrast, genistein-mediated upregulation of CHOP impeded C/EBPβ interaction with the IL-6 promoter, thus disturbing the angiogenic microenvironment. This more malignant microenvironment in EGFRvIII glioblastoma is generated, at least in part, by greater VEGF, IL-6 and C/EBPβ expression. Interaction of C/EBPβ with the IL-6 promoter maintains this angiogenic microenvironment, while disturbance of this dynamically balanced interaction inhibits EGFRvIII tumor proliferation by reducing both VEGF and IL-6 expression.

Borkowski J, Li L, Steinmann U, et al.
Neisseria meningitidis elicits a pro-inflammatory response involving IκBζ in a human blood-cerebrospinal fluid barrier model.
J Neuroinflammation. 2014; 11:163 [PubMed] Article available free on PMC after 01/04/2016 Related Publications
BACKGROUND: The human-specific, Gram-negative bacterium Neisseria meningitidis (Nm) is a leading cause of bacterial meningitis worldwide. The blood-cerebrospinal fluid barrier (BCSFB), which is constituted by the epithelial cells of the choroid plexus (CP), has been suggested as one of the potential entry sites of Nm into the CSF and can contribute to the inflammatory response during infectious diseases of the brain. Toll-like receptors (TLRs) are involved in mediating signal transduction caused by the pathogens.
METHODS: Using a recently established in vitro model of the human BCSFB based on human malignant CP papilloma (HIBCPP) cells we investigated the cellular response of HIBCPP cells challenged with the meningitis-causing Nm strain, MC58, employing transcriptome and RT-PCR analysis, cytokine bead array, and enzyme-linked immunosorbent assay (ELISA). In comparison, we analyzed the answer to the closely related unencapsulated carrier isolate Nm α14. The presence of TLRs in HIBCPP and their role during signal transduction caused by Nm was studied by RT-PCR and the use of specific agonists and mutant bacteria.
RESULTS: We observed a stronger transcriptional response after infection with strain MC58, in particular with its capsule-deficient mutant MC58siaD-, which correlated with bacterial invasion levels. Expression evaluation and Gene Set Enrichment Analysis pointed to a NFκB-mediated pro-inflammatory immune response involving up-regulation of the transcription factor IκBζ. Infected cells secreted significant levels of pro-inflammatory chemokines and cytokines, including, among others, IL8, CXCL1-3, and the IκBζ target gene product IL6. The expression profile of pattern recognition receptors in HIBCPP cells and the response to specific agonists indicates that TLR2/TLR6, rather than TLR4 or TLR2/TLR1, is involved in the cellular reaction following Nm infection.
CONCLUSIONS: Our data show that Nm can initiate a pro-inflammatory response in human CP epithelial cells probably involving TLR2/TLR6 signaling and the transcriptional regulator IκBζ.

Markkula A, Simonsson M, Ingvar C, et al.
IL6 genotype, tumour ER-status, and treatment predicted disease-free survival in a prospective breast cancer cohort.
BMC Cancer. 2014; 14:759 [PubMed] Article available free on PMC after 01/04/2016 Related Publications
BACKGROUND: In breast cancer, high levels of the inflammatory cytokine interleukin-6 (IL-6) have been associated with disease-free survival and treatment resistance. Increased serum levels of IL-6 have been correlated with increased levels of NF-κβ and aromatase expression in adipose tissue. Several IL6 single nucleotide polymorphisms have been associated with breast cancer prognosis, but the impact may differ depending on tumour oestrogen receptor (ER) status. This translational study investigated the association between IL6 genotypes, ER-status, and treatment on the risk of early events among breast cancer patients.
METHODS: The study included 634 25- to 99-year-old primary breast cancer patients in Sweden from 2002-2008. Genotyped IL6 single nucleotide polymorphisms rs1800797, rs1800796, rs1800795, and rs2069849 were analysed separately and as diplotypes. Disease-free survival was assessed for 567 patients. Clinical data, patient-, and tumour-characteristics were obtained from questionnaires, patient charts, population registries, and pathology reports.
RESULTS: The median follow-up time was 5.1 years. IL6 diplotype was not associated with early events for all 567 patients, but AGCC/AGCC diplotype-carriers with ER-negative tumours had an increased risk, (adjusted Hazard Ratio (HR) = 5.91, 95% CI: 1.28-27.42). Any C-carriers (rs1800795) with ER-negative tumours had a higher risk of early events than GG-carriers with ER-negative tumours, (adjusted HR = 3.76, 95% CI: 1.05-13.43), particularly after radiotherapy (adjusted HR = 7.17, 95% CI: 1.16-32.28). Irrespective of ER-status, chemotherapy-treated Any C-carriers had a higher risk of early events than GG-carriers (adjusted HR = 3.42, 95% CI: 1.01-11.54).
CONCLUSIONS: The main finding of the present study was that IL6 genotype was strongly associated with early events among patients with ER-negative tumours, particularly among radiotherapy-treated patients, and among chemotherapy-treated patients irrespective of ER-status. The high risk for early events observed in these subgroups of patients suggests that combined information on IL6 genotype, tumour ER-status, and breast cancer treatment may represent a tool for identifying patients who require more personalised treatment.

Xie GB, Liu WJ, Pan ZJ, et al.
Evolution of the mir-155 family and possible targets in cancers and the immune system.
Asian Pac J Cancer Prev. 2014; 15(18):7547-52 [PubMed] Related Publications
The mir-155 family is not only involved in a diversity of cancers, but also as a regulator of the immune system. However, the evolutionary history of this family is still unclear. The present study indicates that mir-155 evolved independently with lineage-specific gain of miRNAs. In addition, arm switching has occurred in the mir-155 family, and alternative splicing could produce two different lengths of ancestral sequences, implying the alternative splicing can also drive evolution for intragenic miRNAs. Here we screened validated target genes and immunity- related proteins, followed by analyzation of the mir-155 family function by high-throughput methods like the gene ontology (GO) and Kyoto Eneyclopedin of Genes and Genemes (KEGG) pathway enrichment analysis. The high-throughput analysis showed that the CCND1 and EGFR genes were outstanding in being significantly enriched, and the target genes cebpb and VCAM1 and the protein SMAD2 were also vital in mir-155-related immune reponse activities. Therefore, we conclude that the mir-155 family is highly conserved in evolution, and CCND1 and EGFR genes might be potential targets of mir-155 with regard to progress of cancers, while the cebpb and VCAM1 genes and the protein SMAD2 might be key factors in the mir-155 regulated immune activities.

Sheshadri N, Catanzaro JM, Bott AJ, et al.
SCCA1/SERPINB3 promotes oncogenesis and epithelial-mesenchymal transition via the unfolded protein response and IL6 signaling.
Cancer Res. 2014; 74(21):6318-29 [PubMed] Article available free on PMC after 01/11/2015 Related Publications
The serine/cysteine protease inhibitor SCCA1 (SERPINB3) is upregulated in many advanced cancers with poor prognosis, but there is limited information about whether it makes functional contributions to malignancy. Here, we show that SCCA1 expression promoted oncogenic transformation and epithelial-mesenchymal transition (EMT) in mammary epithelial cells, and that SCCA1 silencing in breast cancer cells halted their proliferation. SCCA1 overexpression in neu(+) mammary tumors increased the unfolded protein response (UPR), IL6 expression, and inflammatory phenotypes. Mechanistically, SCCA1 induced a prolonged nonlethal increase in the UPR that was sufficient to activate NF-κB and expression of the protumorigenic cytokine IL6. Overall, our findings established that SCCA1 contributes to tumorigenesis by promoting EMT and a UPR-dependent induction of NF-κB and IL6 autocrine signaling that promotes a protumorigenic inflammation.

Kavarthapu R, Tsai Morris CH, Dufau ML
Prolactin induces up-regulation of its cognate receptor in breast cancer cells via transcriptional activation of its generic promoter by cross-talk between ERα and STAT5.
Oncotarget. 2014; 5(19):9079-91 [PubMed] Article available free on PMC after 01/11/2015 Related Publications
Prolactin (PRL) serves a critical role in breast cancer progression via activation of its cognate receptor. These studies reveal up-regulation of PRLR gene expression by PRL in absence of estradiol in MCF-7 and T47D breast cancer cells. PRL/PRLR via activation of STAT5 that binds a GAS-element in the PRLR gene and the participation of ERα stimulates PRLR transcription/expression. PRL/PRLR induces phosphorylation of ERα through the JAK2/PI3K/MAPK/ERK and JAK2/HER2 activated pathways. The increased recruitment of phospho-ERα, induced by PRL to Sp1 and C/EBPβ at PRLR promoter sites is essential for PRL-induced PRLR transcription. This recruitment is prevented by blockade of PRL expression using RNA interference or ERα phosphorylation using specific inhibitors of PI3K and ERK. Direct evidence is provided for local actions of PRL, independent of estradiol, in the up-regulation of PRLR transcription/expression by an activation-loop between STAT5 and the phospho-ERα/Sp1/C/EBPβ complex with requisite participation of signaling mechanisms. PRL's central role in the up-regulation of PRLR maximizes the action of the endogenous hormone. This study offers mechanistically rational basis for invasiveness fueled by prolactin in refractory states to adjuvant therapies in breast cancer.

Robertson CL, Srivastava J, Siddiq A, et al.
Genetic deletion of AEG-1 prevents hepatocarcinogenesis.
Cancer Res. 2014; 74(21):6184-93 [PubMed] Article available free on PMC after 01/11/2015 Related Publications
Activation of the oncogene AEG-1 (MTDH, LYRIC) has been implicated recently in the development of hepatocellular carcinoma (HCC). In mice, HCC can be initiated by exposure to the carcinogen DEN, which has been shown to rely upon activation of NF-κB in liver macrophages. Because AEG-1 is an essential component of NF-κB activation, we interrogated the susceptibility of mice lacking the AEG-1 gene to DEN-induced hepatocarcinogenesis. AEG-1-deficient mice displayed resistance to DEN-induced HCC and lung metastasis. No difference was observed in the response to growth factor signaling or activation of AKT, ERK, and β-catenin, compared with wild-type control animals. However, AEG-1-deficient hepatocytes and macrophages exhibited a relative defect in NF-κB activation. Mechanistic investigations showed that IL6 production and STAT3 activation, two key mediators of HCC development, were also deficient along with other biologic and epigenetics findings in the tumor microenvironment, confirming that AEG-1 supports an NF-κB-mediated inflammatory state that drives HCC development. Overall, our findings offer in vivo proofs that AEG-1 is essential for NF-κB activation and hepatocarcinogenesis, and they reveal new roles for AEG-1 in shaping the tumor microenvironment for HCC development.

Han JG, Gupta SC, Prasad S, Aggarwal BB
Piperlongumine chemosensitizes tumor cells through interaction with cysteine 179 of IκBα kinase, leading to suppression of NF-κB-regulated gene products.
Mol Cancer Ther. 2014; 13(10):2422-35 [PubMed] Related Publications
Recently, two different reports appeared in prominent journals suggesting a mechanism by which piperlongumine, a pyridine alkaloid, mediates anticancer effects. In the current report, we describe another novel mechanism by which this alkaloid mediates its anticancer effects. We found that piperlongumine blocked NF-κB activated by TNFα and various other cancer promoters. This downregulation was accompanied by inhibition of phosphorylation and degradation of IκBα. Further investigation revealed that this pyridine alkaloid directly interacts with IκBα kinase (IKK) and inhibits its activity. Inhibition of IKK occurred through interaction with its cysteine 179 as the mutation of this residue to alanine abolished the activity of piperlongumine. Inhibition in NF-κB activity downregulated the expression of proteins involved in cell survival (Bcl-2, Bcl-xL, c-IAP-1, c-IAP-2, survivin), proliferation (c-Myc, cyclin D1), inflammation (COX-2, IL6), and invasion (ICAM-1, -9, CXCR-4, VEGF). Overall, our results reveal a novel mechanism by which piperlongumine can exhibit antitumor activity through downmodulation of proinflammatory pathway.

Yu J, Wang Y, Yan F, et al.
Noncanonical NF-κB activation mediates STAT3-stimulated IDO upregulation in myeloid-derived suppressor cells in breast cancer.
J Immunol. 2014; 193(5):2574-86 [PubMed] Related Publications
Immunotherapy for cancer treatment is achieved through the activation of competent immune effector cells and the inhibition of immunosuppressive cells, such as myeloid-derived suppressor cells (MDSCs). Although MDSCs have been shown to contribute to breast cancer development, the mechanism underlying MDSC-mediated immunosuppression is unclear. We have identified a poorly differentiated MDSC subset in breast cancer-suppressing T cell function through STAT3-dependent IDO upregulation. In this study we investigated the mechanisms underlying aberrant expression of IDO in MDSCs. MDSCs were induced by coculturing human CD33(+) myeloid progenitors with MDA-MB-231 breast cancer cells. Increased STAT3 activation in MDSCs was correlated with activation of the noncanonical NF-κB pathway, including increased NF-κB-inducing kinase (NIK) protein level, phosphorylation of cytoplasmic inhibitor of NF-κB kinase α and p100, and RelB-p52 nuclear translocation. Blocking STAT3 activation with the small molecule inhibitor JSI-124 significantly inhibited the accumulation of NIK and IDO expression in MDSCs. Knockdown of NIK in MDSCs suppressed IDO expression but not STAT3 activation. RelB-p52 dimers were found to directly bind to the IDO promoter, leading to IDO expression in MDSCs. IL-6 was found to stimulate STAT3-dependent, NF-κB-mediated IDO upregulation in MDSCs. Furthermore, significant positive correlation between the numbers of pSTAT3(+) MDSCs, IDO(+) MDSCs, and NIK(+) MDSCs was observed in human breast cancers. These results demonstrate a STAT3/NF-κB/IDO pathway in breast cancer-derived MDSCs, which provides insight into understanding immunosuppressive mechanisms of MDSCs in breast cancer.

Li J, Shan F, Xiong G, et al.
EGF-induced C/EBPβ participates in EMT by decreasing the expression of miR-203 in esophageal squamous cell carcinoma cells.
J Cell Sci. 2014; 127(Pt 17):3735-44 [PubMed] Related Publications
Epithelial-mesenchymal transition (EMT) is a developmental program that is associated with esophageal squamous cell carcinoma (ESCC) progression and metastasis. Recently, C/EBPβ has been reported to be an EMT inducer in cancer. However, the detailed molecular mechanisms remain unclear. Here, we report for the first time, that the truncated CCAAT-enhancer-binding protein β (C/EBPβ) LIP isoform is abnormally overexpressed and correlated with cancer metastasis in clinical specimens of human ESCC. Furthermore, we demonstrate that C/EBPβ LIP mediates epithelial growth factor (EGF)-induced EMT and increases migration and invasion of esophageal cancer cells in a manner that is dependent on miR-203 inactivation. Finally, we identified miR-203 as a direct target of C/EBPβ LIP. Disruption of C/EBPβ LIP attenuated the EGF-mediated decrease in miR-203, whereas overexpression of C/EBPβ LIP alone markedly suppressed miR-203. In addition, we demonstrated that C/EBPβ LIP inhibited miR-203 transcription by directly interacting with a conserved distal regulatory element upstream of the miR-203 locus, and in doing so, orchestrated chromatin remodeling. In conclusion, our results have revealed a new regulatory mechanism that involves C/EBPβ-LIP-mediated downregulation of miR-203, which plays a key role in EMT and metastasis.

Liu Q, Boudot A, Ni J, et al.
Cyclin D1 and C/EBPβ LAP1 operate in a common pathway to promote mammary epithelial cell differentiation.
Mol Cell Biol. 2014; 34(16):3168-79 [PubMed] Article available free on PMC after 01/11/2015 Related Publications
Both cyclin D1 and the transcription factor C/EBPβ are required for mammary epithelial cell differentiation; however, the pathway in which they operate is uncertain. Previous analyses of the patterns of gene expression in human tumors suggested a connection between cyclin D1 overexpression and C/EBPβ, but whether this represents a cancer-specific gain of function for cyclin D1 is unknown. C/EBPβ is an intronless gene encoding three protein isoforms--LAP1, LAP2, and LIP. Here, we provide evidence that cyclin D1 engages C/EBPβ in an isoform-specific manner. Cyclin D1 binds to LAP1, an event that activates the transcriptional function of LAP1 by relieving its autoinhibited state effected by intramolecular interactions. Reexpression of LAP1 but not LAP2 or LIP restores the ability of C/EBPβ-deficient mammary epithelial cells to differentiate and does so in a manner dependent on cyclin D1. And cyclin D1-mediated activation of LAP1 participates in mammary epithelial cell differentiation. Our findings indicate that cyclin D1 and C/EBPβ LAP1 operate in a common pathway to promote mammary epithelial cell differentiation.

Xu H, Lai W, Zhang Y, et al.
Tumor-associated macrophage-derived IL-6 and IL-8 enhance invasive activity of LoVo cells induced by PRL-3 in a KCNN4 channel-dependent manner.
BMC Cancer. 2014; 14:330 [PubMed] Article available free on PMC after 01/11/2015 Related Publications
BACKGROUND: Tumor-associated macrophages (TAMs) are known to promote cancer progression and metastasis through the release of a variety of cytokines. Phosphatase of regenerating liver (PRL-3) has been considered as a marker of colorectal cancer (CRC) liver metastasis. Our previous research suggests that PRL-3 can enhance the metastasis of CRC through the up-regulation of intermediate-conductance Ca2+-activated K+ (KCNN4) channel, which is dependent on the autocrine secretion of tumor necrosis factor-alpha (TNF-α). However, whether TAMs participate in the progression and metastasis of CRC induced by PRL-3 remains unknown.
METHODS: We used flow cytometry, coculture, western blotting, invasion assays, real-time quantitative PCR, chromatin immunoprecipitation, luciferase reporter assays, and immunofluorescence staining to determine the effect of TAMs on the ability of PRL-3 to promote invasiveness of CRC cells.
RESULTS: In this study, we found that TAMs facilitated the metastasis of CRC induced by PRL-3. When TAMs were cocultured with CRC cells, the expression of KCNN4 was increased in TAMs and the invasion of CRC cells was enhanced. Furthermore, cytokines that were secreted by TAMs, such as IL-6 and IL-8, were also significantly increased. This response was attenuated by treating TAMs with the KCNN4 channel-specific inhibitor, 1-[(2-chlorophenyl) diphenylmethyl]-1H-pyrazole (TRAM-34), which suggested that KCNN4 channels may be involved in inducing the secretion of IL-6 and IL-8 by TAMs and improving CRC cell invasiveness. Moreover, the expression of KCNN4 channels in TAMs was regulated through the NF-κB signal pathway, which is activated by TNF-α from CRC cells. Immunofluorescence analysis of colorectal specimens indicated that IL-6 and IL-8 double positive cells in the stroma showed positive staining for the TAM marker CD68, suggesting that TAMs produce IL-6 and IL-8. Increased numbers of these cells correlated with higher clinical stage.
CONCLUSIONS: Our findings suggested that TAMs participate in the metastasis of CRC induced by PRL-3 through the TNF-α mediated secretion of IL-6 and IL-8 in a paracrine manner.

Li P, Harris D, Liu Z, et al.
STAT3-activated GM-CSFRα translocates to the nucleus and protects CLL cells from apoptosis.
Mol Cancer Res. 2014; 12(9):1267-82 [PubMed] Article available free on PMC after 01/09/2015 Related Publications
UNLABELLED: Here, it was determined that chronic lymphocytic leukemia (CLL) cells express the α subunit, but not the β subunit, of the granulocyte-macrophage colony-stimulating factor receptor (GM-CSFR/CSF2R). GM-CSFRα was detected on the surface, in the cytosol, and in the nucleus of CLL cells via confocal microscopy, cell fractionation, and GM-CSFRα antibody epitope mapping. Because STAT3 is frequently activated in CLL and the GM-CSFRα promoter harbors putative STAT3 consensus binding sites, MM1 cells were transfected with truncated forms of the GM-CSFRα promoter, then stimulated with IL6 to activate STAT3 and to identify STAT3-binding sites. Chromatin immunoprecipitation (ChIP) and an electoromobility shift assay (EMSA) confirmed STAT3 occupancy to those promoter regions in both IL6-stimulated MM1 and CLL cells. Transfection of MM1 cells with STAT3-siRNA or CLL cells with STAT3-shRNA significantly downregulated GM-CSFRα mRNA and protein levels. RNA transcripts, involved in regulating cell survival pathways, and the proteins KAP1 (TRIM28) and ISG15 coimmunoprecipitated with GM-CSFRα. GM-CSFRα-bound KAP1 enhanced the transcriptional activity of STAT3, whereas GM-CSFRα-bound ISG15 inhibited the NF-κB pathway. Nevertheless, overexpression of GM-CSFRα protected MM1 cells from dexamethasone-induced apoptosis, and GM-CSFRα knockdown induced apoptosis in CLL cells, suggesting that GM-CSFRα provides a ligand-independent survival advantage.
IMPLICATIONS: Constitutively, activation of STAT3 induces the expression of GM-CSFRα that protects CLL cells from apoptosis, suggesting that inhibition of STAT3 or GM-CSFRα may benefit patients with CLL.

Jeffries SJ, Jones L, Harrison CJ, Russell LJ
IGH@ translocations co-exist with other primary rearrangements in B-cell precursor acute lymphoblastic leukemia.
Haematologica. 2014; 99(8):1334-42 [PubMed] Article available free on PMC after 01/09/2015 Related Publications
Primary established genetic abnormalities in B-cell precursor acute lymphoblastic leukemia include high hyperdiploidy (51-65 chromosomes), the translocations t(12;21)(p13;q22)/ETV6-RUNX1 fusion and t(9;22)(q34;q11)/BCR-ABL1 fusion, MLL rearrangements and intrachromosomal amplification of chromosome 21. These rearrangements are of prognostic and therapeutic relevance and are usually mutually exclusive. We identified 28 patients at diagnosis with both a primary genetic rearrangement and an immunoglobulin heavy chain locus translocation using chromosomal analysis and fluorescence in situ hybridization. Among these patients, the immunoglobulin heavy chain locus translocation partner gene was identified in six (CRLF2, CEBPA, CEBPB, TRA/D@, IGF2BP1 and IGK@). Clonal architecture was investigated in 17 patients using multiple color interphase fluorescence in situ hybridization analysis, which showed that the translocation was acquired as a secondary abnormality in ten patients, in four patients the etiology was undetermined and in three patients it was observed in a separate clone from the primary chromosomal rearrangement. These findings demonstrate the co-existence of immunoglobulin heavy chain locus translocations with other primary chromosomal rearrangements either in the same or separate clones, which may have prognostic significance in B-cell precursor acute lymphoblastic leukemia. Clinical trials: UKALLXII: Study ID n. ISRCTN77346223 and ALL2003: Study ID n. ISRCTN07355119.

Campion CG, Labrie M, Grosset AA, St-Pierre Y
The CCAAT/enhancer-binding protein beta-2 isoform (CEBPβ-2) upregulates galectin-7 expression in human breast cancer cells.
PLoS One. 2014; 9(5):e95087 [PubMed] Article available free on PMC after 01/09/2015 Related Publications
Galectin-7 is considered a gene under the control of p53. However, elevated expression of galectin-7 has been reported in several forms of cancer harboring an inactive p53 pathway. This is especially true for breast cancer where galectin-7 expression is readily expressed in a high proportion in basal-like breast cancer tissues, conferring cancer cells with increased resistance to cell death and metastatic properties. These observations suggest that other transcription factors are capable of inducing galectin-7 expression. In the present work, we have examined the role of CCAAT/enhancer-binding protein beta (C/EBPβ) in inducing expression of galectin-7. C/EBP proteins have been shown to contribute to breast cancer by upregulating pro-metastatic genes. We paid particular attention to C/EBPβ-2 (also known as LAP2), the most transcriptionally active of the C/EBPβ isoforms. Our results showed that ectopic expression of C/EBPβ-2 in human breast cancer cells was sufficient to induce expression of galectin-7 at both the mRNA and protein levels. In silico analysis further revealed the presence of an established CEBP element in the galectin-7 promoter. Mutation of this binding site abolished the transcriptional activity of the galectin-7 promoter. Chromatin immunoprecipitation analysis confirmed that C/EBPβ-2 binds to the endogenous galectin-7 promoter. Analysis of galectin-7 protein expression in normal epithelia and in breast carcinoma by immunohistochemistry further showed the expression pattern of C/EBPβ closely micmicked that of galectin-7, most notably in mammary myoepithelial cells and basal-like breast cancer where galectin-7 is preferentially expressed. Taken together, our findings suggest that C/EBPβ is an important mediator of galectin-7 gene activation in breast cancer cells and highlight the different transcriptional mechanisms controlling galectin-7 in cancer cells.

Cho HY, Lee SW
TLR5 activation by flagellin induces doxorubicin resistance via interleukin-6 (IL-6) expression in two multiple myeloma cells.
Cell Immunol. 2014 May-Jun; 289(1-2):27-35 [PubMed] Related Publications
Multiple myeloma (MM) is an incurable B-cell hematologic malignancy characterized by the clonal expansion of malignant plasma cells in the bone marrow (BM). MM cells interact with various cells within the BM microenvironment, leading to skeletal destruction, angiogenesis, and drug resistance. Therefore, control of the cell-host interaction and growth factors is important to improve patient outcome with conventional treatments. In this study, we investigated flagellin-induced cell proliferation, cytokines expression, and the mechanisms of cancer drug resistance that lead to the failure of cytotoxic therapies for MM. The human MM line KMS28BM expresses the TLR5 gene as well as the protein at high levels. When treated with the specific TLR5 ligand flagellin, KMS28BM cells showed increased proliferation, increased IgG λ production, and high-level expression and secretion of the pro-inflammatory cytokine IL-6, via NF-κB activation through p38 and PI3K/AKT signaling. Furthermore, flagellin-stimulated KMS28BM cells were shown to have "increased doxorubicin and apoptosis resistance" through the inhibition of caspases and PARP activity, and this result was reversed by blocking IL-6. Thus, increased cell viability and the chemoresistance of flagellin-stimulated KMS28BM cells may result from autocrine or paracrine signaling mediated by secreted IL-6. These findings indicate that TLR5 activation by flagellin may elicit chemoresistance in MM patients who have suffered from recurrent bacterial infections.

Chiang KC, Tsui KH, Chung LC, et al.
Celastrol blocks interleukin-6 gene expression via downregulation of NF-κB in prostate carcinoma cells.
PLoS One. 2014; 9(3):e93151 [PubMed] Article available free on PMC after 01/09/2015 Related Publications
Interleukin-6 (IL-6), a multifunctional cytokine, contributes to proliferation or differentiation of prostate carcinoma cells in a highly cell type-specific manner. Celastrol (3-hydroxy-24-nor-2oxo-1(10),3,5,7-friedelatetrane-29-oic acid), also named as tripterine, is extracted from root of Chinese traditional herb Tripterygiumwilfordii Hook f with potent anti-inflammatory and anti-cancer activities. In this study, we evaluated the molecular mechanisms of celastrol on cell proliferation and IL-6 gene expression in prostate carcinoma cells. 3H-thymidine incorporation and flow cytometric analysis indicated that celastrol treatments arrested the cell cycle at the G0/G1 phase, thus attenuating cell proliferation in prostate carcinoma PC-3 cells; moreover, celastrol induced cell apoptosis at higher dosage. Knockdown of IL-6 attenuated the anti-proliferative effect of celastrol on PC-3 cells. Results from ELISA and 5'-deletion transient gene expression assays indicated that celastrol treatment decreased IL-6 secretion and gene expression, and this effect is dependent on the NF-κB response element within IL-6 promoter area since mutation of the NF-κB response element from AAATGTCCCATTTTCCC to AAATGTTACATTTTCCC by site-directed mutagenesis abolished the inhibition of celastrol on the IL-6 promoter activity. Celastrol also attenuated the activation of PMA and TNFα on the gene expression and secretion of IL-6 in PC-3 cells. Immunoblot assays revealed that celastrol treatment downregulated the expressions of IKKα, p50 and p65, supporting the 5'-deletion transient gene expression assay result that celastrol blocked IL-6 expression through the NF-κB pathway in PC-3 cells. For the first time, our results concluded that celastrol attenuates PC-3 cell proliferation via downregulation of IL-6 gene expression through the NF-κB-dependent pathway.

Li J, Shan F, Xiong G, et al.
Transcriptional regulation of miR-146b by C/EBPβ LAP2 in esophageal cancer cells.
Biochem Biophys Res Commun. 2014; 446(1):267-71 [PubMed] Related Publications
Recent clinical study indicated that up-regulation of miR-146b was associated with poor overall survival of patients in esophageal squamous cell carcinoma. However, the underlying mechanism of miR-146b dysregulation remains to be explored. Here we report that miR-146b promotes cell proliferation and inhibits cell apoptosis in esophageal cancer cell lines. Mechanismly, two C/EBPβ binding motifs are located in the miR-146b promoter conserved region. Among the three isoforms of C/EBPβ, C/EBPβ LAP2 positively regulated miR-146b expression and increases miR-146b levels in a dose-dependent manner through transcription activation of miR-146b gene. Together, these results suggest a miR-146b regulatory mechanism involving C/EBPβ, which may contribute to the up-regulation of miR-146b in esophageal squamous cell carcinoma.

Gérard C, Gonze D, Lemaigre F, Novák B
A model for the epigenetic switch linking inflammation to cell transformation: deterministic and stochastic approaches.
PLoS Comput Biol. 2014; 10(1):e1003455 [PubMed] Article available free on PMC after 01/09/2015 Related Publications
Recently, a molecular pathway linking inflammation to cell transformation has been discovered. This molecular pathway rests on a positive inflammatory feedback loop between NF-κB, Lin28, Let-7 microRNA and IL6, which leads to an epigenetic switch allowing cell transformation. A transient activation of an inflammatory signal, mediated by the oncoprotein Src, activates NF-κB, which elicits the expression of Lin28. Lin28 decreases the expression of Let-7 microRNA, which results in higher level of IL6 than achieved directly by NF-κB. In turn, IL6 can promote NF-κB activation. Finally, IL6 also elicits the synthesis of STAT3, which is a crucial activator for cell transformation. Here, we propose a computational model to account for the dynamical behavior of this positive inflammatory feedback loop. By means of a deterministic model, we show that an irreversible bistable switch between a transformed and a non-transformed state of the cell is at the core of the dynamical behavior of the positive feedback loop linking inflammation to cell transformation. The model indicates that inhibitors (tumor suppressors) or activators (oncogenes) of this positive feedback loop regulate the occurrence of the epigenetic switch by modulating the threshold of inflammatory signal (Src) needed to promote cell transformation. Both stochastic simulations and deterministic simulations of a heterogeneous cell population suggest that random fluctuations (due to molecular noise or cell-to-cell variability) are able to trigger cell transformation. Moreover, the model predicts that oncogenes/tumor suppressors respectively decrease/increase the robustness of the non-transformed state of the cell towards random fluctuations. Finally, the model accounts for the potential effect of competing endogenous RNAs, ceRNAs, on the dynamics of the epigenetic switch. Depending on their microRNA targets, the model predicts that ceRNAs could act as oncogenes or tumor suppressors by regulating the occurrence of cell transformation.

Shi WY, Liu KD, Xu SG, et al.
Gene expression analysis of lung cancer.
Eur Rev Med Pharmacol Sci. 2014; 18(2):217-28 [PubMed] Related Publications
OBJECTIVE: We aim to explore the expression difference between lung cancer cells and normal lung cells, and to investigate the mechanism of lung cancer development. Besides, we predicted the potential target site of transcriptional factors and microRNAs for differentially expressed genes (DEGs), which may help to regulate expression of DEGs. Small molecules were also identified to cure lung cancer.
MATERIALS AND METHODS: Gene expression profiles we used were downloaded from Gene Expression Omnibus (GEO) using accession number of GSE2378. Firstly, we identified differential genes between lung cancer cells and normal lung cells by using R package limma. Then, we detected the processes and pathways that changed in lung cancer cells by Gene Ontology (GO) and KEGG pathway enrichment analysis. Potential target sites of transcriptional factors and microRNAs were also detected based on gene annotation data in MSigDB. Finally, small molecule drugs were screened via querying Connectivity Map database.
RESULTS: We obtained 2961 differentially expressed genes between lung cancer cells and normal lung cells. Besides changes in cell cycle, metabolic processes and proteasome were also dramatically disordered. Some DEGs shared target sites of the transcription factor such as E2F, ETS and CEBPB. Target sites of hsa-miR-196a and hsa-miR-200c were also significantly enriched by DEGs. Iloprost simulated the state of normal cells, while MS-275 might be potential pathogenic substances.
CONCLUSIONS: We investigate the lung cancer from Gene Ontology, pathway, transcription factors and microRNAs based on gene expression profiles. All these results may facilitate lung cancer treatment with a new breakthrough.

Xiang M, Birkbak NJ, Vafaizadeh V, et al.
STAT3 induction of miR-146b forms a feedback loop to inhibit the NF-κB to IL-6 signaling axis and STAT3-driven cancer phenotypes.
Sci Signal. 2014; 7(310):ra11 [PubMed] Article available free on PMC after 01/09/2015 Related Publications
Interleukin-6 (IL-6)-mediated activation of signal transducer and activator of transcription 3 (STAT3) is a mechanism by which chronic inflammation can contribute to cancer and is a common oncogenic event. We discovered a pathway, the loss of which is associated with persistent STAT3 activation in human cancer. We found that the gene encoding the tumor suppressor microRNA miR-146b is a direct STAT3 target gene, and its expression was increased in normal breast epithelial cells but decreased in tumor cells. Methylation of the miR-146b promoter, which inhibited STAT3-mediated induction of expression, was increased in primary breast cancers. Moreover, we found that miR-146b inhibited nuclear factor κB (NF-κB)-dependent production of IL-6, subsequent STAT3 activation, and IL-6/STAT3-driven migration and invasion in breast cancer cells, thereby establishing a negative feedback loop. In addition, higher expression of miR-146b was positively correlated with patient survival in breast cancer subtypes with increased IL6 expression and STAT3 phosphorylation. Our results identify an epigenetic mechanism of crosstalk between STAT3 and NF-κB relevant to constitutive STAT3 activation in malignancy and the role of inflammation in oncogenesis.

Singh R, Sharma MC, Sarkar C, et al.
Transcription factor C/EBP-β mediates downregulation of dipeptidyl-peptidase III expression by interleukin-6 in human glioblastoma cells.
FEBS J. 2014; 281(6):1629-41 [PubMed] Related Publications
Dipeptidyl-peptidase III (DPP III) is a cytosolic metallo-aminopeptidase implicated in various physiological and pathological processes. A previous study from our laboratory indicated an elevated expression of DPP III in glioblastoma (U87MG) cells. In the present study we investigated the role of interleukin-6 (IL-6), a pleiotropic cytokine produced by glial tumors, in the regulation of DPP III expression. Immunohistochemistry, western blotting and quantitative RT-PCR were used for quantitation of DPP III and IL-6 in human glioblastoma cells and tumors. Cell transfections and DPP III promoter reporter assays were performed to study the transcriptional regulation of DPP III by IL-6. Promoter deletion analysis, site directed mutagenesis, chromatin immunoprecipitation assays and small interfering RNA (siRNA) technology was employed to elucidate the molecular mechanism of IL-6 mediated regulation of DPP III expression in glioblastoma cells. Our results for the first time demonstrate a negative correlation (r = 0.632, P = 0.01) between DPP III and IL-6 in both human tumors and cultured glioblastoma cells. Treatment of U87MG cells with IL-6 significantly decreased DPP III expression with a concomitant increase in the levels of transcription factor CCAAT/enhancer binding protein beta (C/EBP-β). Deletion/mutagenesis of C/EBP-β binding motif of DPP III promoter significantly increased its activity and abolished its responsiveness to IL-6. This effect could also be mimicked by C/EBP-β siRNA. In conclusion our study for the first time demonstrates C/EBP-β mediated transcriptional downregulation of DPP III by IL-6. Our results demonstrating a negative correlation between IL-6 and DPP III taken together with the previously reported prognostic significance of this cytokine in glioblastoma suggests that DPP III may prove useful as a prognostic marker.

Kling P, Modig C, Mujahed H, et al.
Differential regulation of the rainbow trout (Oncorhynchus mykiss) MT-A gene by nuclear factor interleukin-6 and activator protein-1.
BMC Mol Biol. 2013; 14:28 [PubMed] Article available free on PMC after 01/09/2015 Related Publications
BACKGROUND: Previously we have identified a distal region of the rainbow trout (Oncorhynchus mykiss) metallothionein-A (rtMT-A) enhancer region, being essential for free radical activation of the rtMT-A gene. The distal promoter region included four activator protein 1 (AP1) cis-acting elements and a single nuclear factor interleukin-6 (NF-IL6) element. In the present study we used the rainbow trout hepatoma (RTH-149) cell line to further examine the involvement of NF-IL6 and AP1 in rtMT-A gene expression following exposure to oxidative stress and tumour promotion.
RESULTS: Using enhancer deletion studies we observed strong paraquat (PQ)-induced rtMT-A activation via NF-IL6 while the AP1 cis-elements showed a weak but significant activation. In contrast to mammals the metal responsive elements were not activated by oxidative stress. Electrophoretic mobility shift assay (EMSA) mutation analysis revealed that the two most proximal AP1 elements, AP11,2, exhibited strong binding to the AP1 consensus sequence, while the more distal AP1 elements, AP13,4 were ineffective. Phorbol-12-myristate-13-acetate (PMA), a known tumor promoter, resulted in a robust induction of rtMT-A via the AP1 elements alone. To determine the conservation of regulatory functions we transfected human Hep G2 cells with the rtMT-A enhancer constructs and were able to demonstrate that the cis-elements were functionally conserved. The importance of NF-IL6 in regulation of teleost MT is supported by the conservation of these elements in MT genes from different teleosts. In addition, PMA and PQ injection of rainbow trout resulted in increased hepatic rtMT-A mRNA levels.
CONCLUSIONS: These studies suggest that AP1 primarily is involved in PMA regulation of the rtMT-A gene while NF-IL6 is involved in free radical regulation. Taken together this study demonstrates the functionality of the NF-IL6 and AP-1 elements and suggests an involvement of MT in protection during pathological processes such as inflammation and cancer.

Verdelli D, Nobili L, Todoerti K, et al.
Molecular events underlying interleukin-6 independence in a subclone of the CMA-03 multiple myeloma cell line.
Genes Chromosomes Cancer. 2014; 53(2):154-67 [PubMed] Related Publications
We explored the molecular mechanisms involved in the establishement of CMA-03/06, an IL-6-independent variant of the multiple myeloma cell line CMA-03 previously generated in our Institution. CMA-03/06 cells grow in the absence of IL-6 with a doubling time comparable with that of CMA-03 cells; neither the addition of IL6 (IL-6) to the culture medium nor co-culture with multipotent mesenchymal stromal cells increases the proliferation rate, although they maintain the responsiveness to IL-6 stimulation as demonstrated by STAT1, STAT3, and STAT5 induction. IL-6 independence of CMA-03/06 cells is not apparently due to the development of an autocrine IL-6 loop, nor to the observed moderate constitutive activation of STAT5 and STAT3, since STAT3 silencing does not affect cell viability or proliferation. When compared to the parental cell line, CMA-03/06 cells showed an activated pattern of the NF-κB pathway. This finding is supported by gene expression profiling (GEP) analysis identifying an appreciable fraction of modulated genes (28/308) in the CMA-03/06 subclone reported to be involved in this pathway. Furthermore, although more resistant to apoptotic stimuli compared to the parental cell line, CMA-03/06 cells display a higher sensibility to NF-κB inhibition induced by bortezomib. Finally, GEP analysis suggests an involvement of a number of cytokines, which might contribute to IL-6 independence of CMA-03/06 by stimulating growth and antiapoptotic processes. In conclusion, the parental cell-line CMA-03 and its variant CMA-03/06 represent a suitable model to further investigate molecular mechanisms involved in the IL-6-independent growth of myeloma cells.

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