Targeted Disruption of Hotair Leads to Homeotic Transformation and Gene Derepression
2013; 5 (1): 3-12
HOXB13 Mediates Tamoxifen Resistance and Invasiveness in Human Breast Cancer by Suppressing ERa and Inducing IL-6 Expression.
2013; 73 (17): 5449-5458
Long noncoding RNAs (lncRNAs) are thought to be prevalent regulators of gene expression, but the consequences of lncRNA inactivation in vivo are mostly unknown. Here, we show that targeted deletion of mouse Hotair lncRNA leads to derepression of hundreds of genes, resulting in homeotic transformation of the spine and malformation of metacarpal-carpal bones. RNA sequencing and conditional inactivation reveal an ongoing requirement of Hotair to repress HoxD genes and several imprinted loci such as Dlk1-Meg3 and Igf2-H19 without affecting imprinting choice. Hotair binds to both Polycomb repressive complex 2, which methylates histone H3 at lysine 27 (H3K27), and Lsd1 complex, which demethylates histone H3 at lysine 4 (H3K4) in vivo. Hotair inactivation causes H3K4me3 gain and, to a lesser extent, H3K27me3 loss at target genes. These results reveal the function and mechanisms of Hotair lncRNA in enforcing a silent chromatin state at Hox and additional genes.
View details for DOI 10.1016/j.celrep.2013.09.003
View details for Web of Science ID 000326152100002
A long noncoding RNA maintains active chromatin to coordinate homeotic gene expression
2011; 472 (7341): 120-U158
Most breast cancers expressing the estrogen receptor α (ERα) are treated successfully with the receptor antagonist tamoxifen (TAM), but many of these tumors recur. Elevated expression of the homeodomain transcription factor HOXB13 correlates with TAM-resistance in ERα-positive (ER+) breast cancer, but little is known regarding the underlying mechanism. Our comprehensive evaluation of HOX gene expression using tiling microarrays, with validation, showed that distant metastases from TAM-resistant patients also displayed high HOXB13 expression, suggesting a role for HOXB13 in tumor dissemination and survival. Here we show that HOXB13 confers TAM resistance by directly downregulating ERα transcription and protein expression. HOXB13 elevation promoted cell proliferation in vitro and growth of tumor xenografts in vivo. Mechanistic investigations showed that HOXB13 transcriptionally upregulated interleukin (IL)-6, activating the mTOR pathway via STAT3 phosphorylation to promote cell proliferation and fibroblast recruitment. Accordingly, mTOR inhibition suppressed fibroblast recruitment and proliferation of HOXB13-expressing ER+ breast cancer cells and tumor xenografts, alone or in combination with TAM. Taken together, our results establish a function for HOXB13 in TAM resistance through direct suppression of ERα and they identify the IL-6 pathways as mediator of disease progression and recurrence. Cancer Res; 73(17); 5449-58. ©2013 AACR.
View details for DOI 10.1158/0008-5472.CAN-13-1178
View details for PubMedID 23832664
Long non-coding RNA HOTAIR reprograms chromatin state to promote cancer metastasis
2010; 464 (7291): 1071-U148
The genome is extensively transcribed into long intergenic noncoding RNAs (lincRNAs), many of which are implicated in gene silencing. Potential roles of lincRNAs in gene activation are much less understood. Development and homeostasis require coordinate regulation of neighbouring genes through a process termed locus control. Some locus control elements and enhancers transcribe lincRNAs, hinting at possible roles in long-range control. In vertebrates, 39 Hox genes, encoding homeodomain transcription factors critical for positional identity, are clustered in four chromosomal loci; the Hox genes are expressed in nested anterior-posterior and proximal-distal patterns colinear with their genomic position from 3' to 5'of the cluster. Here we identify HOTTIP, a lincRNA transcribed from the 5' tip of the HOXA locus that coordinates the activation of several 5' HOXA genes in vivo. Chromosomal looping brings HOTTIP into close proximity to its target genes. HOTTIP RNA binds the adaptor protein WDR5 directly and targets WDR5/MLL complexes across HOXA, driving histone H3 lysine 4 trimethylation and gene transcription. Induced proximity is necessary and sufficient for HOTTIP RNA activation of its target genes. Thus, by serving as key intermediates that transmit information from higher order chromosomal looping into chromatin modifications, lincRNAs may organize chromatin domains to coordinate long-range gene activation.
View details for DOI 10.1038/nature09819
View details for Web of Science ID 000289199400049
View details for PubMedID 21423168
Localized scleroderma and systemic sclerosis: Is there a connection?
BEST PRACTICE & RESEARCH IN CLINICAL RHEUMATOLOGY
2007; 21 (6): 1025-1036
Large intervening non-coding RNAs (lincRNAs) are pervasively transcribed in the genome yet their potential involvement in human disease is not well understood. Recent studies of dosage compensation, imprinting, and homeotic gene expression suggest that individual lincRNAs can function as the interface between DNA and specific chromatin remodelling activities. Here we show that lincRNAs in the HOX loci become systematically dysregulated during breast cancer progression. The lincRNA termed HOTAIR is increased in expression in primary breast tumours and metastases, and HOTAIR expression level in primary tumours is a powerful predictor of eventual metastasis and death. Enforced expression of HOTAIR in epithelial cancer cells induced genome-wide re-targeting of Polycomb repressive complex 2 (PRC2) to an occupancy pattern more resembling embryonic fibroblasts, leading to altered histone H3 lysine 27 methylation, gene expression, and increased cancer invasiveness and metastasis in a manner dependent on PRC2. Conversely, loss of HOTAIR can inhibit cancer invasiveness, particularly in cells that possess excessive PRC2 activity. These findings indicate that lincRNAs have active roles in modulating the cancer epigenome and may be important targets for cancer diagnosis and therapy.
View details for DOI 10.1038/nature08975
View details for Web of Science ID 000276635000045
View details for PubMedID 20393566
Activation of nuclear hormone receptor peroxisome proliferator-activated receptor-delta accelerates intestinal adenoma growth
2004; 10 (3): 245-247
Excess fibrosis of the skin is a clinical hallmark of both localized scleroderma and systemic sclerosis. Localized scleroderma is generally thought to be a skin-limited disease whereas systemic sclerosis can have a wide range of internal organ involvement. Recent data suggest that a subset of patients with juvenile localized scleroderma can go on to develop systemic involvement of their disease. This raises the question of what the connection is, if any, between localized scleroderma and systemic sclerosis.
View details for DOI 10.1016/j.berh.2007.09.003
View details for Web of Science ID 000252093900005
View details for PubMedID 18068859
Peroxisome proliferator-activated receptor gamma-mediated differentiation - A mutation in colon cancer cells reveals divergent and cell type-specific mechanisms
JOURNAL OF BIOLOGICAL CHEMISTRY
2003; 278 (25): 22669-22677
We treated Apc(min) mice, which are predisposed to intestinal polyposis, with a selective synthetic agonist of peroxisome proliferator-activated receptor-delta (PPAR-delta). Exposure of Apc(min) mice to the PPAR-delta ligand GW501516 resulted in a significant increase in the number and size of intestinal polyps. The most prominent effect was on polyp size; mice treated with the PPAR-delta activator had a fivefold increase in the number of polyps larger than 2 mm. Our results implicate PPAR-delta in the regulation of intestinal adenoma growth.
View details for DOI 10.1038/nm993
View details for Web of Science ID 000189297700034
View details for PubMedID 14758356
Cyclooxygenase-1 is overexpressed and promotes angiogenic growth factor production in ovarian cancer
2003; 63 (5): 906-911
Activation of the nuclear hormone receptor peroxisome proliferator-activated receptor gamma (PPARgamma) inhibits cell growth and induces differentiation in both adipocyte and epithelial cell lineages, although it is unclear whether this occurs through common or cell-type specific mechanisms. We have identified four human colon cancer cell lines that do no undergo growth inhibition or induce markers of differentiation after exposure to PPARgamma agonists. Sequence analysis of the PPARgamma gene revealed that all four cell lines contain a previously unidentified point mutation in the ninth alpha-helix of the ligand binding domain at codon 422 (K422Q). The mutant receptor did not exhibit any defects in DNA binding or retinoid X receptor heterodimerization and was transcriptionally active in an artificial reporter assay. However, only retroviral transduction of the wild-type (WT), but not mutant, receptor could restore PPARgamma ligand-induced growth inhibition and differentiation in resistant colon cancer cell lines. In contrast, there was no difference in the ability of fibroblast cells expressing WT or K422Q mutant receptor to undergo growth inhibition, express adipocyte differentiation markers, or uptake lipid after treatment with a PPARgamma agonist. Finally, analysis of direct PPARgamma target genes in colon cancer cells expressing the WT or K422Q mutant allele suggests that the mutation may disrupt the ability of PPARgamma to repress the basal expression of a subset of genes in the absence of exogenous ligand. Collectively, these data argue that codon 422 may be a part of a co-factor(s) interaction domain necessary for PPARgamma to induce terminal differentiation in epithelial, but not adipocyte, cell lineages and argues that the receptor induces growth inhibition and differentiation via cell lineage-specific mechanisms.
View details for DOI 10.1074/jbc.M300637200
View details for Web of Science ID 000183503900067
View details for PubMedID 12591919
Expression of hypoxia-inducible factors in the peri-implantation mouse uterus is regulated in a cell-specific and ovarian steroid hormone-dependent manner - Evidence for differential function of HIFs during early pregnancy
JOURNAL OF BIOLOGICAL CHEMISTRY
2003; 278 (9): 7683-7691
Inhibition of cyclooygenase-2 (COX-2) catalytic activity has proven successful in restricting the growth of epithelial-derived cancers in vivo. Whether COX-2 inhibitor therapy would be beneficial in the prevention and/or treatment of ovarian cancer, the most lethal gynecological malignancy worldwide, is not known. Most patients with ovarian cancer undergo cytoreductive therapy. Because many of the cytotoxic drugs used to treat ovarian cancer induce COX-2 expression, samples from patients that had not undergone cytoreductive therapy were specifically chosen for COX isoform expression analysis. A majority of specimens exhibited elevated levels of COX-1, not COX-2, mRNA, and protein compared with normal ovarian tissue. Focal regions within the tumor expressing high COX-1 also had elevated levels of pro-angiogenic proteins. Selective inhibition of COX-1, not COX-2, inhibited arachidonic acid-stimulated vascular endothelial growth factor production, which could be reversed by cotreatment with prostaglandin E(2). Thus, COX-1 may contribute to carcinoma development in the ovary through stimulation of neovascularization. Clinical studies testing the efficacy of COX inhibition as adjuvant therapy for ovarian cancer may see more beneficial effects with adjuvant therapy with either a COX-1 selective or nonselective cyclooxygenase inhibitor as compared with a COX-2 selective drug.
View details for Web of Science ID 000181327700003
View details for PubMedID 12615701
Peroxisome proliferator-activated receptor gamma and transforming growth factor-beta pathways inhibit intestinal epithelial cell growth by regulating levels of TSC-22
JOURNAL OF BIOLOGICAL CHEMISTRY
2003; 278 (9): 7431-7438
Increased uterine vascular permeability and angiogenesis are hallmarks of implantation and placentation. These events are profoundly influenced by vascular endothelial growth factor (VEGF). We previously showed that VEGF isoforms and VEGF receptors are expressed in the uterus, suggesting the role of VEGF in uterine vascular permeability and angiogenesis required for implantation and decidualization. We have recently shown that estrogen promotes uterine vascular permeability but inhibits angiogenesis, whereas progesterone stimulates angiogenesis with little effect on vascular permeability. However, the mechanism of differential steroid hormonal regulation of uterine angiogenesis remains unresolved. Oxygen homeostasis is essential for cell survival and is primarily mediated by hypoxia-inducible factors (HIFs). These factors are intimately associated with vascular events and induce VEGF expression by binding to the hypoxia response element in the VEGF promoter. HIFalpha isoforms function by forming heterodimers with the aryl hydrocarbon nuclear translocator (ARNT) (HIF-beta) family members. There is very limited information on the relationship among HIFs, ARNTs, and VEGF in the uterus during early pregnancy, although the role of HIFs in regulating VEGF and angiogenesis in cancers is well documented. Using molecular and physiological approaches, we here show that uterine expression of HIFs and ARNTs does not correlate with VEGF expression during the preimplantation period (days 1-4) in mice. In contrast, their expression follows the localization of uterine VEGF expression with increasing angiogenesis during the postimplantation period (days 5-8). This disparate pattern of uterine HIFs, ARNTs, and VEGF expression on days 1-4 of pregnancy suggests HIFs have multiple roles in addition to the regulation of angiogenesis during the peri-implantation period. Using pharmacological, molecular, and genetic approaches, we also observed that although progesterone primarily up-regulates uterine HIF-1alpha expression, estrogen transiently stimulates that of HIF-2alpha.
View details for DOI 10.1074/jbc.M211390200
View details for Web of Science ID 000181195100133
View details for PubMedID 12482866
Colorectal cancer prevention and treatment by inhibition of cyclooxygenase-2
NATURE REVIEWS CANCER
2001; 1 (1): 11-21
Peroxisome proliferator-activated receptor gamma (PPARgamma) and transforming growth factor-beta (TGF-beta) are key regulators of epithelial cell biology. However, the molecular mechanisms by which either pathway induces growth inhibition and differentiation are incompletely understood. We have identified transforming growth factor-simulated clone-22 (TSC-22) as a target gene of both pathways in intestinal epithelial cells. TSC-22 is member of a family of leucine zipper containing transcription factors with repressor activity. Although little is known regarding its function in mammals, the Drosophila homolog of TSC-22, bunched, plays an essential role in fly development. The ability of PPARgamma to induce TSC-22 was not dependent on an intact TGF-beta1 signaling pathway and was specific for the gamma isoform. Localization studies revealed that TSC-22 mRNA is enriched in the postmitotic epithelial compartment of the normal human colon. Cells transfected with wild-type TSC-22 exhibited reduced growth rates and increased levels of p21 compared with vector-transfected cells. Furthermore, transfection with a dominant negative TSC-22 in which both repressor domains were deleted was able to reverse the p21 induction and growth inhibition caused by activation of either the PPARgamma or TGF-beta pathways. These results place TSC-22 as an important downstream component of PPARgamma and TGF-beta signaling during intestinal epithelial cell differentiation.
View details for DOI 10.1074/jbc.M208076200
View details for Web of Science ID 000181195100103
View details for PubMedID 12468551
Activation of peroxisome proliferator-activated receptor gamma suppresses nuclear factor kappa B-mediated apoptosis induced by Helicobacter pylori in gastric epithelial cells
JOURNAL OF BIOLOGICAL CHEMISTRY
2001; 276 (33): 31059-31066
Population-based studies have established that long-term intake of non-steroidal anti-inflammatory drugs (NSAIDs), compounds that inhibit the enzymatic activity of cyclooxygenase (COX), reduces the relative risk for developing colorectal cancer. These studies led to the identification of a molecular target, COX-2, that is involved in tumour promotion during colorectal cancer progression. Recent studies in humans indicate that therapy with specific COX-2 inhibitors might be an effective approach to colorectal cancer prevention and treatment.
View details for Web of Science ID 000180397000010
View details for PubMedID 11900248
Target genes of peroxisome proliferator-activated receptor gamma in colorectal cancer cells
JOURNAL OF BIOLOGICAL CHEMISTRY
2001; 276 (32): 29681-29687
Helicobacter pylori colonization leads to epithelial cell hyperproliferation within inflamed mucosa, but levels of apoptosis vary, suggesting that imbalances between rates of cell production and loss may contribute to differences in gastric cancer risk among infected populations. Peroxisome proliferator-activated receptor gamma (PPARgamma) regulates inflammatory and growth responses of intestinal epithelial cells. We determined whether activation of PPARgamma modified H. pylori-induced apoptosis in gastric epithelial cells. PPARgamma was expressed and functionally active in gastric epithelial cell lines sensitive to H. pylori-induced apoptosis. PPARgamma ligands 15d-PGJ(2) and BRL-49653 significantly attenuated H. pylomicronri-induced apoptosis, effects that could be reversed by co-treatment with a specific PPARgamma antagonist. Cyclopentanone prostaglandins that do not bind and activate PPARgamma had no effects on H. pylori-induced apoptosis. The ability of H. pylori to activate nuclear factor (NF)-kappaB and increase levels of the NF-kappaB target IL-8 was blocked by co-treatment with PPARgamma agonists, and direct inhibition of NF-kappaB also abolished H. pylori-stimulated apoptosis. These results suggest that activation of the PPARgamma pathway attenuates the ability of H. pylori to induce NF-kappaB-mediated apoptosis in gastric epithelial cells. Because PPARgamma regulates a multitude of host responses, activation of this receptor may contribute to varying levels of cellular turnover as well as the diverse pathologic outcomes associated with chronic H. pylori colonization.
View details for Web of Science ID 000170472900063
View details for PubMedID 11397815
15S-hydroxyeicosatetraenoic acid activates peroxisome proliferator-activated receptor gamma and inhibits proliferation in PC3 prostate carcinoma cells
2001; 61 (2): 497-503
Activation of the nuclear hormone peroxisome proliferator-activated receptor gamma (PPARgamma) inhibits cell growth and promotes differentiation in a broad spectrum of epithelial derived tumor cell lines. Here we utilized microarray technology to identify PPARgamma gene targets in intestinal epithelial cells. For each gene, the induction or repression was seen with two structurally distinct PPARgamma agonists, and the change in expression could be blocked by co-treatment with a specific PPARgamma antagonist. A majority of the genes could be regulated independently by a retinoid X receptor specific agonist. Genes implicated in lipid transport or storage (adipophilin and liver fatty acid-binding protein) were also activated by agonists of PPAR subtypes alpha and/or delta. In contrast, PPARgamma-selective targets included genes linked to growth regulatory pathways (regenerating gene IA), colon epithelial cell maturation (GOB-4 and keratin 20), and immune modulation (neutrophil-gelatinase-associated lipocalin). Additionally, three different genes of the carcinoembryonic antigen family were induced by PPARgamma. Cultured cells treated with PPARgamma ligands demonstrated an increase in Ca(2+)-independent, carcinoembryonic antigen-dependent homotypic aggregation, suggesting a potential role for PPARgamma in regulating intercellular adhesion. Collectively, these results will help define the mechanisms by which PPARgamma regulates intestinal epithelial cell biology.
View details for Web of Science ID 000170558000013
View details for PubMedID 11397807
Prostacyclin-mediated activation of peroxisome proliferator-activated receptor delta in colorectal cancer
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
2000; 97 (24): 13275-13280
15-Lipoxygenase (15-LOX)-2 is expressed in benign prostate secretory cells and benign prostate produces 15S-hydroxyeicosatetraenoic acid (15S-HETE) from exogenous arachidonic acid (AA). In contrast, 15S-LOX-2 and 15S-HETE formation are reduced in prostate carcinoma (Pca). The mechanisms whereby reduced 15-LOX-2 may contribute to Pca development or progression are not known. We investigated the expression of peroxisome proliferator-activated receptor (PPAR) gamma in benign and malignant prostate tissues and the ability of 15S-HETE to activate PPARgamma-dependent transcription and modulate proliferation of the Pca cell line PC3. In contrast to benign prostate and similar to most Pca tissues, 15-LOX-2 mRNA was not detected in PC3 cells, and they did not produce detectable 15-HETE from [14C]AA. By reverse transcription-PCR, PPARgamma mRNA was present in 18 of 18 benign and 9 of 9 tumor specimens. The PPARgamma ligand BRL 49653 and 15S-HETE caused a dose-dependent inhibition of PC3 proliferation in a 14-day soft agar colony-forming assay (IC50 of 3 and 30 microM, respectively). 15S-HETE (10 microM) caused greater inhibition than 10 microM 15R-HETE. At 3 days, BRL 49653 and 15S-HETE caused a slight increase in cells in G0-G1 and a corresponding decrease in cells in S phase. In PC3 cells transiently transfected with a luciferase reporter linked to a PPAR response element, 1 microM BRL 49653 and 10 microM 15S-HETE caused approximately threefold and greater than twofold induction of PPAR-dependent transcription, respectively. By quantitative real-time reverse transcription-PCR and Northern analysis, 3-day treatment with BRL 49653 and 15S-HETE caused a reduction of PPARgamma expression but a marked up-regulation of the PPAR response element containing adipocyte type fatty acid binding protein. These results support the hypothesis that 15-LOX-2-derived 15S-HETE may constitute an endogenous ligand for PPARgamma in the prostate and that loss of this pathway by reduced expression of 15-LOX-2 may contribute to increased proliferation and reduced differentiation in prostate carcinoma.
View details for Web of Science ID 000166819800019
View details for PubMedID 11212240
Activation of PPAR gamma leads to inhibition of anchorage-independent growth of human colorectal cancer cells
1998; 115 (5): 1049-1055
There is evidence from both genetic and pharmacologic studies to suggest that the cyclooxygenase-2 (COX-2) enzyme plays a causal role in the development of colorectal cancer. However, little is known about the identity or role of the eicosanoid receptor pathways activated by COX-derived prostaglandins (PG). We previously have reported that COX-2-derived prostacyclin promotes embryo implantation in the mouse uterus via activation of the nuclear hormone receptor peroxisome proliferator-activated receptor (PPAR) delta. In light of the recent finding that PPARdelta is a target of beta-catenin transactivation, it is important to determine whether this signaling pathway is operative during the development of colorectal cancer. Analysis of PPARdelta mRNA in matched normal and tumor samples revealed that expression of PPARdelta, similar to COX-2, is up-regulated in colorectal carcinomas. In situ hybridization studies demonstrate that PPARdelta is expressed in normal colon and localized to the epithelial cells at the very tips of the mucosal glands. In contrast, expression of PPARdelta mRNA in colorectal tumors was more widespread with increased levels in transformed epithelial cells. Analysis of PPARdelta and COX-2 mRNA in serial sections suggested they were colocalized to the same region within a tumor. Finally, transient transfection assays established that endogenously synthesized prostacyclin (PGI(2)) could serve as a ligand for PPARdelta. In addition, the stable PGI(2) analog, carbaprostacyclin, and a synthetic PPARdelta agonist induced transactivation of endogenous PPARdelta in human colon carcinoma cells. We conclude from these observations that PPARdelta, similar to COX-2, is aberrantly expressed in colorectal tumors and that endogenous PPARdelta is transcriptionally responsive to PGI(2). However, the functional consequence of PPARdelta activation in colon carcinogenesis still needs to be determined.
View details for Web of Science ID 000165476300069
View details for PubMedID 11087869
Peroxisomal proliferator-activated receptor gamma (PPARgamma) is a nuclear hormone receptor that provides a direct link between fatty acid metabolism and control of gene transcription. The objective of this study was to determine the biological effect(s) of PPARgamma activation in colorectal carcinoma cells.PPARgamma expression and activity were measured in 4 human colon cancer cell lines using reverse-transcription polymerase chain reaction, immunoprecipitation and immunoblotting, and transient reporter gene assays. The effects of activated PPARgamma in these cell lines were assessed in cellular proliferation and anchorage-independent growth assays. Flow cytometry was used to determine the effects of PPARgamma activation on progression through the cell cycle.PPARgamma was expressed in all 4 colon cancer cell lines examined and was transcriptionally functional in 3 of the 4. Treatment of these cells with a selective PPARgamma activator (BRL 49653) resulted in inhibition of anchorage-independent growth. The degree of growth inhibition correlated with the level of functional PPARgamma present. Finally, activation of PPARgamma resulted in G1 cell cycle arrest.Activation of the PPARgamma pathway in colon cancer cells has potent antiproliferative effects, suggesting that this nuclear hormone receptor may provide a novel target for prevention and treatment of colorectal cancer in humans.
View details for Web of Science ID 000076664900007
View details for PubMedID 9797355