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The Kruppel-like factor 9 (KLF9) network in HEC-1-A endometrial carcinoma cells suggests the carcinogenic potential of dys-regulated KLF9 expression.

ABSTRACT: Krüppel-like factor 9 (KLF9) is a transcriptional regulator of uterine endometrial cell proliferation, adhesion and differentiation; processes essential for pregnancy success and which are subverted during tumorigenesis. The network of endometrial genes controlled by KLF9 is largely unknown. Over-expression of KLF9 in the human endometrial cancer cell line HEC-1-A alters cell morphology, proliferative indices, and differentiation, when compared to KLF9 under-expressing HEC-1-A cells. This cell line provides a unique model for identifying KLF9 downstream gene targets and signaling pathways.HEC-1-A sub-lines differing in relative levels of KLF9 were subjected to microarray analysis to identify differentially-regulated RNAs.KLF9 under-expression induced twenty four genes. The KLF9-suppressed mRNAs encode protein participants in: aldehyde metabolism (AKR7A2, ALDH1A1); regulation of the actin cytoskeleton and cell motility (e.g., ANK3, ITGB8); cellular detoxification (SULT1A1, ABCC4); cellular signaling (e.g., ACBD3, FZD5, RAB25, CALB1); and transcriptional regulation (PAX2, STAT1). Sixty mRNAs were more abundant in KLF9 over-expressing sub-lines. The KLF9-induced mRNAs encode proteins which participate in: regulation and function of the actin cytoskeleton (COTL1, FSCN1, FXYD5, MYO10); cell adhesion, extracellular matrix and basement membrane formation (e.g., AMIGO2, COL4A1, COL4A2, LAMC2, NID2); transport (CLIC4); cellular signaling (e.g., BCAR3, MAPKAPK3); transcriptional regulation [e.g., KLF4, NR3C1 (glucocorticoid receptor), RXRalpha], growth factor/cytokine actions (SLPI, BDNF); and membrane-associated proteins and receptors (e.g., CXCR4, PTCH1). In addition, the abundance of mRNAs that encode hypothetical proteins (KLF9-inhibited: C12orf29 and C1orf186; KLF9-induced: C10orf38 and C9orf167) were altered by KLF9 expression. Human endometrial tumors of high tumor grade had decreased KLF9 mRNA abundance.KLF9 influences the expression of uterine epithelial genes through mechanisms likely involving its transcriptional activator and repressor functions and which may underlie altered tumor biology with aberrant KLF9 expression.

SUBMITTER: Simmen FA

PROVIDER: S-EPMC2542371 | biostudies-literature | 2008 Sep

REPOSITORIES: biostudies-literature

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The Krüppel-like factor 9 (KLF9) network in HEC-1-A endometrial carcinoma cells suggests the carcinogenic potential of dys-regulated KLF9 expression.

Simmen Frank A FA Su Ying Y Xiao Rijin R Zeng Zhaoyang Z Simmen Rosalia C M RC

Reproductive biology and endocrinology : RB&E 20080910

<h4>Background</h4>Krüppel-like factor 9 (KLF9) is a transcriptional regulator of uterine endometrial cell proliferation, adhesion and differentiation; processes essential for pregnancy success and which are subverted during tumorigenesis. The network of endometrial genes controlled by KLF9 is largely unknown. Over-expression of KLF9 in the human endometrial cancer cell line HEC-1-A alters cell morphology, proliferative indices, and differentiation, when compared to KLF9 under-expressing HEC-1-A ...[more]

PMID: 18783612

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Project description:The hypothalamic peptide gonadotropin inhibitory hormone (GnIH) is a relatively novel hypothalamic neuropeptide, identified in 2000. It can influence the hypothalamic-pituitary-gonadal axis and reproductive function through various neuroendocrine systems. The present study aimed to explore the effects and potential underlying molecular mechanism of RFamide-related peptide-3 (RFRP-3) injection on the uterine fluid protein profile of ovariectomized estrogen-primed (OEP) rats using proteomics. In addition, the possible effects of RFRP-3 on the viability and apoptosis of the human endometrial cancer cell line HEC-1A and associated molecular mechanism were investigated. The OEP rat model was established through injection with GnIH/RFRP-3 through the lateral ventricle. At 6 h after injection, the protein components of uterine fluid of rats in the experimental and control groups were analyzed using liquid chromatography (LC)-tandem mass spectrometry (MS/MS). Differentially expressed proteins (DEPs) were analyzed using the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. Protein-protein interactions (PPI) were investigated using the STRING database. PPI networks were then established before hub proteins were selected using OmicsBean software. The expression of one of the hub proteins, Kras, was then detected using western blot analysis. Cell Counting Kit-8, Annexin V-FITC/PI, reverse transcription-quantitative PCR and western blotting were also performed to analyze cell viability and apoptosis. In total, 417 DEPs were obtained using LC-MS/MS, including 279 upregulated and 138 downregulated proteins. GO analysis revealed that the majority of the DEPs were secretory proteins. According to KEGG enrichment analysis, the DEPs found were generally involved in tumor-associated pathways. In particular, five hub proteins, namely G protein subunit α (Gna)13, Gnaq, Gnai3, Kras and MMP9, were obtained following PPI network analysis. Western blot analysis showed that expression of the hub protein Kras was downregulated following treatment with 10,000 ng/ml RFRP-3. RFRP-3 treatment (10,000 ng/ml) also suppressed HEC-1A cell viability, induced apoptosis, downregulated Bcl-2 and upregulated Bax protein expression, compared with those in the control group. In addition, compared with those in the control group, RFRP-3 significantly reduced the mRNA expression levels of PI3K, AKT and mTOR, while upregulating those of LC3-II. Compared with those in the control group, RFRP-3 significantly decreased the protein expression levels of PI3K, AKT, mTOR and p62, in addition to decreasing AKT phosphorylation. By contrast, RFRP-3 significantly increased the LC3-II/I ratio and G protein-coupled receptor 147 (GPR147) protein expression. In conclusion, the present data suggest that RFRP-3 can alter the protein expression profile of the uterine fluid of OEP rats by upregulating MMP9 expression whilst downregulating that of key hub proteins Gna13, GnaQ, Gnai3 and Kras. Furthermore, RFRP-3 can inhibit HEC-1A cell viability while promoting apoptosis. The underlying molecular mechanism may involve activation of GPR147 receptor by the direct binding of RFRP-3, which further downregulates the hub protein Kras to switch on the PI3K/AKT/mTOR pathway. This subsequently reduces the Bcl-2 expression and promotes Bax expression to induce autophagy.

Project description:BackgroundCircadian disruption is an emerging driver of breast cancer (BCa), with epidemiological studies linking shift work and chronic jet lag to increased BCa risk. Indeed, several clock genes participate in the gating of mitotic entry, regulation of DNA damage response, and epithelial-to-mesenchymal transition, thus impacting BCa etiology. Dysregulated estrogen (17β-estradiol, E2) and glucocorticoid (GC) signaling prevalent in BCa may further contribute to clock desynchrony by directly regulating the expression and cycling dynamics of genes comprising the local breast oscillator. In this study, we investigated the tumor suppressor gene, Krüppel-like factor 9 (KLF9), as an important point of crosstalk between hormone signaling and the circadian molecular network, and further examine its functional role in BCa.MethodsThrough meta-analysis of publicly available RNA- and ChIP-sequencing datasets from BCa tumor samples and cell lines, and gene expression analysis by RT-qPCR and enhancer- reporter assays, we elucidated the molecular mechanism behind the clock and hormone regulation of KLF9. Lentiviral knockdown and overexpression of KLF9 in three distinct breast epithelial cell lines (MCF10A, MCF7 and MDA-MB-231) was generated to demonstrate the role of KLF9 in orthogonal assays on breast epithelial survival, proliferation, apoptosis, and migration.ResultsWe determined that KLF9 is a direct GC receptor target in mammary epithelial cells, and that induction is likely mediated through coordinate transcriptional activation from multiple GC-responsive enhancers in the KLF9 locus. More interestingly, rhythmic expression of KLF9 in MCF10A cells was abolished in the highly aggressive MDA-MB-231 line. In turn, forced expression of KLF9 altered the baseline and GC/E2-responsive expression of several clock genes, indicating that KLF9 may function as a regulator of the core clock machinery. Characterization of the role of KLF9 using complementary cancer hallmark assays in the context of the hormone-circadian axis revealed that KLF9 plays a tumor-suppressive role in BCa regardless of molecular subtype. KLF9 potentiated the anti-tumorigenic effects of GC in E2 receptor + luminal MCF7 cells, while it restrained GC-enhanced oncogenicity in triple-negative MCF10A and MDA-MB-231 cells.ConclusionsTaken together, our findings support that dysregulation of KLF9 expression and oscillation in BCa impinges on circadian network dynamics, thus ultimately affecting the BCa oncogenic landscape.

Project description:IntroductionA large body of evidence has established a pattern of altered functioning in the immune system, autonomic nervous system and hypothalamic pituitary adrenal axis in chronic fatigue syndrome. However, the relationship between components within and between these systems is unclear. In this paper we investigated the underlying network structure of the autonomic system in patients and controls, and a larger network comprising all three systems in patients alone.MethodsIn a sample of patients and controls we took several measures of autonomic nervous system output during 10 minutes of supine rest covering tests of blood pressure variability, heart rate variability and cardiac output. Awakening salivary cortisol was measured on each of two days with participants receiving 0.5mg dexamethasone during the afternoon of the first day. Basal plasma cytokine levels and the in vitro cytokine response to dexamethasone were also measured. Symptom outcome measures used were the fatigue impact scale and cognitive failures questionnaire. Mutual information criteria were used to construct networks describing the dependency amongst variables. Data from 42 patients and 9 controls were used in constructing autonomic networks, and 15 patients in constructing the combined network.ResultsThe autonomic network in patients showed a more uneven distribution of information, with two distinct modules emerging dominated by systolic blood pressure during active stand and end diastolic volume and stroke volume respectively. The combined network revealed strong links between elements of each of the three regulatory systems, characterised by three higher modules the centres of which were systolic blood pressure during active stand, stroke volume and ejection fraction respectively.ConclusionsCFS is a complex condition affecting physiological systems. It is important that novel analytical techniques are used to understand the abnormalities that lead to CFS. The underlying network structure of the autonomic system is significantly different to that of controls, with a small number of individual nodes being highly influential. The combined network suggests links across regulatory systems which shows how alterations in single nodes might spread throughout the network to produce alterations in other, even distant, nodes. Replication in a larger cohort is warranted.

Dataset Information

The Kruppel-like factor 9 (KLF9) network in HEC-1-A endometrial carcinoma cells suggests the carcinogenic potential of dys-regulated KLF9 expression.

Publications

The Krüppel-like factor 9 (KLF9) network in HEC-1-A endometrial carcinoma cells suggests the carcinogenic potential of dys-regulated KLF9 expression.

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