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Togo Ikuta,
Yasuhito Kobayashi,
Masato Kitazawa,
Kazuhiro Shiizaki,
Naoki Itano,
Tetsuo Noda,
Sven Pettersson, Lorenz Poellinger,
Yoshiaki Fujii-Kuriyama,
Shun'ichiro Taniguchi,
Kaname Kawajiri
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ABSTRACT: The aryl hydrocarbon receptor (AhR) plays a suppressive role in cecal carcinogenesis by CUL4B/AhR-mediated ubiquitylation and degradation of β-catenin, which is activated by xenobiotics and natural ligands. AhR-deficient (AhR-/-) mice develop cecal tumors with severe inflammation. To elucidate whether the tumors develop autonomously in AhR-/- mice due to impaired β-catenin degradation or in association with accelerated inflammation, we performed two kinds of experiments using germ-free (GF) AhR-/- mice and compound mutant mice lacking genes for AhR and ASC which plays an essential role in caspase-1 activation in inflammasomes. Both GF AhR-/- and AhR-/-•ASC-/- mice showed considerably reduced tumor development compared with that in AhR-/- mice albeit in a "cancer-prone" state with aberrant β-catenin accumulation. Blocking of the IL-1β signaling pathway by treatment with a caspase-1 inhibitor, YVAD reduced cecal tumorigenesis in AhR-/- mice. STAT3 activation was detected in the cecal epithelium of the AhR-/- mice due to enhanced IL-6 production. An inhibitor of the STAT3 signaling pathway, AG490 suppressed the tumor formation. ASC-mediated inflammation was also found to play a critical role in tumor development in ApcMin/+ mice, a mouse model of familial adenomatous polyposis. Collectively, these results revealed an important role of the bacteria-triggered or ASC-mediated inflammation signaling pathway in the intestinal tumorigenesis of mice and suggest a possible chemical therapeutic intervention, including AhR-ligands and inhibitors of the inflammation pathway.
Carcinogenesis 03/2013; · 5.70 Impact Factor
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Fredrik Lanner,
Kian Leong Lee,
Germán Camargo Ortega,
Marcus Sohl,
Xiujuan Li,
Shaobo Jin,
Emil M Hansson,
Lena Claesson-Welsh, Lorenz Poellinger,
Urban Lendahl,
Filip Farnebo
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ABSTRACT: Hypoxia (low oxygen) and Notch signaling are two important regulators of vascular development, but how they interact in controlling the choice between arterial and venous fates for endothelial cells during vasculogenesis is less well understood. In this report, we show that hypoxia and Notch signaling intersect in promotion of arterial differentiation. Hypoxia upregulated expression of the Notch ligand Dll4 and increased Notch signaling, in a process requiring the vasoactive hormone adrenomedullin. Notch signaling also upregulated Dll4 expression, leading to a positive feedback loop sustaining Dll4 expression and Notch signaling. In addition, hypoxia-mediated upregulation of the arterial marker genes Depp, connexin40 (Gja5), Cxcr4 and Hey1 required Notch signaling. In conclusion, the data reveal an intricate interaction between hypoxia and Notch signaling in the control of endothelial cell differentiation, including a hypoxia/adrenomedullin/Dll4 axis that initiates Notch signaling and a requirement for Notch signaling to effectuate hypoxia-mediated induction of the arterial differentiation program.
Stem cells and development 02/2013; · 4.15 Impact Factor
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ABSTRACT: The Aryl hydrocarbon Receptor (AhR) is a signal regulated transcription factor, which is canonically activated by the direct binding of xenobiotics. In addition, switching cells from adherent to suspension culture also activates the AhR, representing a non-xenobiotic, physiological activation of AhR signaling. Here, we show that the AhR is recruited to target gene enhancers in both ligand (YH439) treated and suspension cells, suggesting a common mechanism of target gene induction between these two routes of AhR activation. However, gene expression profiles critically differ between xenobiotic and suspension activated AhR signaling. Por and Cldnd1 were regulated predominately by ligand treatments, while in contrast, ApoER2 and Ganc were regulated predominately by the suspension condition. Classic xenobiotic metabolizing AhR targets such as Cyp1a1, Cyp1b1, and Nqo1 were regulated by both ligand and suspension conditions. Temporal expression patterns of AhR target genes were also found to vary, with examples of transient activation, transient repression, or sustained alterations in expression. Furthermore, sequence analysis coupled with ChIP assays and reporter gene analysis identified a functional XRE (xenobiotic response element) in the intron1 of mouse Tiparp gene, which was also bound by HIF-1α during hypoxia, and features a concatemer of 4 XRE cores (GCGTG). Our data suggest that this XRE concatemer site concurrently regulates the expression of both Tiparp gene and its cis anti-sense non-coding RNA following ligand or suspension induced AhR activation. This work lends novel insights into how AhR signaling drives different transcriptional programs via the ligand versus suspension modes of activation.
Molecular pharmacology 08/2012; · 4.53 Impact Factor
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ABSTRACT: The dioxin/aryl hydrocarbon receptor (AhR) mediates most toxic effects of dioxins. In utero/lactational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) impairs fetal/neonatal development and the developing male reproductive tract are among the most sensitive tissues. TCDD causes antiestrogenic responses in rodent mammary gland and uterus and in human breast cancer cell lines in the presence of estrogen. Also, more recently an estrogen-like effect of TCDD/AhR has been suggested in the absence of estrogen. A transgenic mouse expressing a constitutively active AhR (CA-AhR) was developed as a model mimicking a situation of constant exposure to AhR agonists. Male and female reproductive tissues of CA-AhR mice were characterized for some of the effects commonly seen after dioxin exposure. Sexually mature CA-AhR female mice showed decreased uterus weight, while an uterotrophic assay in immature CA-AhR mice resulted in increased uterus weight. In immature mice, both TCDD-exposure and CA-AhR increased the expression of the estrogen receptor target gene Cathepsin D. When co-treated with 17β-estradiol no increase in Cathepsin D levels occurred in either TCDD-exposed or CA-AhR mice. In sexually mature male CA-AhR mice the weights of testis and ventral prostate were decreased and the epididymal sperm reserve was reduced. The results of the present study are in accordance with previous studies on dioxin-exposed rodents in that an activated AhR (here CA-AhR) leads to antiestrogenic effects in the presence of estrogen, but to estrogenic effects in the absence of estrogen. These results suggest the CA-AhR mouse model as a useful tool for studies of continuous low activity of the AhR from early development, resembling the human exposure situation.
Chemosphere 12/2011; 85(11):1701-6. · 3.21 Impact Factor
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ABSTRACT: Nodal and Activin are morphogens of the TGFbeta superfamily of signaling molecules that direct differential cell fate decisions in a dose- and distance-dependent manner. During early embryonic development the Nodal/Activin pathway is responsible for the specification of mesoderm, endoderm, node, and mesendoderm. In contradiction to this drive towards cellular differentiation, the pathway also plays important roles in the maintenance of self-renewal and pluripotency in embryonic and epiblast stem cells. The molecular basis behind stem cell interpretation of Nodal/Activin signaling gradients and the undertaking of disparate cell fate decisions remains poorly understood. Here, we show that any perturbation of endogenous signaling levels in mouse embryonic stem cells leads to their exit from self-renewal towards divergent differentiation programs. Increasing Nodal signals above basal levels by direct stimulation with Activin promotes differentiation towards the mesendodermal lineages while repression of signaling with the specific Nodal/Activin receptor inhibitor SB431542 induces trophectodermal differentiation. To address how quantitative Nodal/Activin signals are translated qualitatively into distinct cell fates decisions, we performed chromatin immunoprecipitation of phospho-Smad2, the primary downstream transcriptional factor of the Nodal/Activin pathway, followed by massively parallel sequencing, and show that phospho-Smad2 binds to and regulates distinct subsets of target genes in a dose-dependent manner. Crucially, Nodal/Activin signaling directly controls the Oct4 master regulator of pluripotency by graded phospho-Smad2 binding in the promoter region. Hence stem cells interpret and carry out differential Nodal/Activin signaling instructions via a corresponding gradient of Smad2 phosphorylation that selectively titrates self-renewal against alternative differentiation programs by direct regulation of distinct target gene subsets and Oct4 expression.
PLoS Genetics 06/2011; 7(6):e1002130. · 8.69 Impact Factor
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Heather Main,
Kian Leong Lee,
Henry Yang,
Saija Haapa-Paananen,
Henrik Edgren,
Shaobo Jin,
Cecilia Sahlgren,
Olli Kallioniemi, Lorenz Poellinger,
Bing Lim,
Urban Lendahl
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ABSTRACT: Interaction between key signaling mechanisms is important to generate the diversity in signaling output required for proper control of cellular differentiation and function, although the molecular manifestations of such cross-talk are only partially understood. Notch signaling and the cellular response to hypoxia intersect at different points in the signaling cascades, and in this report we analyze the consequences of this cross-talk at the transcriptome level. Mouse ES cells were subjected to various combinations of hypoxia and/or activated Notch signaling, and the transcriptome changes could be grouped into different categories, reflecting various modes of hypoxia and Notch signaling integration. Two principal categories of novel Notch- and hypoxia-induced genes were identified: (i) a larger set of Notch or hypoxic target genes which were induced by one pathway and not significantly affected by the activity status of the other pathway and (ii) a smaller set of genes co-regulated by Notch and hypoxia. In the latter category, we identified genes that were induced by hypoxia and the expression of which was enhanced by active Notch signaling and another group of genes that were induced by Notch and hypoxia independently. Several of the hypoxia- and Notch-induced genes were found to be upregulated in various forms of cancer. Identification of genes co-regulated by the two pathways may provide a molecular platform to better understand the intersection between the two signaling cascades in normal development and cancer.
Experimental Cell Research 05/2010; 316(9):1610-24. · 3.58 Impact Factor
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Na Zhang,
Zhenxing Fu,
Sarah Linke,
Johana Chicher,
Jeffrey J Gorman,
DeeAnn Visk,
Gabriel G Haddad, Lorenz Poellinger,
Daniel J Peet,
Frank Powell,
Randall S Johnson
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ABSTRACT: Factor inhibiting HIF-1alpha (FIH) is an asparaginyl hydroxylase. Hydroxylation of HIF-alpha proteins by FIH blocks association of HIFs with the transcriptional coactivators CBP/p300, thus inhibiting transcriptional activation. We have created mice with a null mutation in the FIH gene and found that it has little or no discernable role in mice in altering classical aspects of HIF function, e.g., angiogenesis, erythropoiesis, or development. Rather, it is an essential regulator of metabolism: mice lacking FIH exhibit reduced body weight, elevated metabolic rate, hyperventilation, and improved glucose and lipid homeostasis and are resistant to high-fat-diet-induced weight gain and hepatic steatosis. Neuron-specific loss of FIH phenocopied some of the major metabolic phenotypes of the global null animals: those mice have reduced body weight, increased metabolic rate, and enhanced insulin sensitivity and are also protected against high-fat-diet-induced weight gain. These results demonstrate that FIH acts to a significant degree through the nervous system to regulate metabolism.
Cell metabolism 05/2010; 11(5):364-78. · 17.35 Impact Factor
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ABSTRACT: Embryonic stem (ES) cells continuously decide whether to maintain pluripotency or differentiate. While exogenous leukemia inhibitory factor and BMP4 perpetuate a pluripotent state, less is known about the factors initiating differentiation. We show that heparan sulfate (HS) proteoglycans are critical coreceptors for signals inducing ES cell differentiation. Genetic targeting of NDST1 and NDST2, two enzymes required for N-sulfation of proteoglycans, blocked differentiation. This phenotype was rescued by HS presented in trans or by soluble heparin. NaClO, which reduces sulfation of proteoglycans, potently blocked differentiation of wild-type cells. Mechanistically, N-sulfation was identified to be critical for functional autocrine fibroblast growth factor 4 (FGF4) signaling. Microarray analysis identified the pluripotency maintaining transcription factors Nanog, KLF2/4/8, Tbx3, and Tcf3 to be negatively regulated, whereas markers of differentiation such as Gbx2, Dnmt3b, FGF5, and Brachyury were induced by sulfation-dependent FGF receptor (FGFR) signaling. We show that several of these genes are heterogeneously expressed in ES cells, and that targeting of heparan sulfation or FGFR-signaling facilitated a homogenous Nanog/KLF4/Tbx3 positive ES cell state. This finding suggests that the recently discovered heterogeneous state of ES cells is regulated by HS-dependent FGFR signaling. Similarly, culturing blastocysts with NaClO eliminated GATA6-positive primitive endoderm progenitors generating a homogenous Nanog-positive inner cell mass. Functionally, reduction of sulfation robustly improved de novo ES cell derivation efficiency. We conclude that N-sulfated HS is required for FGF4 signaling to maintain ES cells primed for differentiation in a heterogeneous state. Inhibiting this pathway facilitates a more naïve ground state. STEM CELLS 2010;28:191–200
Stem Cells 11/2009; 28(2):191 - 200. · 7.78 Impact Factor
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ABSTRACT: The sensing of oxygen levels and maintenance of oxygen homeostasis is crucial for cells. The hypoxic-sensitive regulation of gene expression allows information about the oxygen status to be converted into appropriate cellular responses. Although there is a core transcriptional pathway, the signalling cascade can be modified to allow diversity and specificity in the transcriptional output. In this Review, we discuss recent advances in our understanding of the mechanisms and factors that contribute to the observed diversity and specificity. A deeper knowledge about how hypoxic signalling is tuned will further our understanding of the cellular hypoxic response in normal physiology and how it becomes derailed in disease.
Nature Reviews Genetics 11/2009; 10(12):821-32. · 38.08 Impact Factor
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ABSTRACT: Activation of transcription in response to low oxygen tension is mediated by the hypoxia-inducible factor-1 (HIF-1). HIF-1 is a heterodimer of two proteins: aryl hydrocarbon receptor nuclear translocator and the oxygen-regulated HIF-1 alpha. The C-terminal activation domain of HIF-1 alpha has been shown to interact with cysteine/histidine-rich region 1 (CH1) of the coactivator CBP/p300 in a hypoxia-dependent manner. However, HIF forms lacking C-terminal activation domain (naturally occurring or genetically engineered) are still able to activate transcription of target genes in hypoxia. Here, we demonstrate that the N-terminal activation domain (N-TAD) of HIF-1 alpha interacts with endogenous CBP and that this interaction facilitates its transactivation function. Our results show that interaction of HIF-1 alpha N-TAD with CBP/p300 is mediated by the CH3 region of CBP known to interact with, among other factors, p53. Using fluorescence resonance energy transfer experiments, we demonstrate that N-TAD interacts with CH3 in vivo. Coimmunoprecipitation assays using endogenous proteins showed that immunoprecipitation of CBP in hypoxia results in the recovery of a larger fraction of HIF-1 alpha than of p53. Chromatin immunoprecipitation demonstrated that at 1% O(2) CBP is recruited to a HIF-1 alpha but not to a p53 target gene. Upon activation of both pathways, lower levels of chromatin-associated CBP were detected at either target gene promoter. These results identify CBP as the coactivator directly interacting with HIF-1 alpha N-TAD and mediating the transactivation function of this domain. Thus, we suggest that in hypoxia HIF-1 alpha is a major CBP-interacting transcription factor that may compete with other CBP-dependent factors, including p53, for limiting amounts of this coactivator, underscoring the complexity in the regulation of gene expression by HIF-1 alpha.
Journal of Biological Chemistry 11/2009; 285(4):2601-9. · 4.77 Impact Factor
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Kaname Kawajiri,
Yasuhito Kobayashi,
Fumiaki Ohtake,
Togo Ikuta,
Yoshibumi Matsushima,
Junsei Mimura,
Sven Pettersson,
Richard S Pollenz,
Toshiyuki Sakaki,
Takatsugu Hirokawa,
Tetsu Akiyama,
Masafumi Kurosumi, Lorenz Poellinger,
Shigeaki Kato,
Yoshiaki Fujii-Kuriyama
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ABSTRACT: Intestinal cancer is one of the most common human cancers. Aberrant activation of the canonical Wnt signaling cascade, for example, caused by adenomatous polyposis coli (APC) gene mutations, leads to increased stabilization and accumulation of beta-catenin, resulting in initiation of intestinal carcinogenesis. The aryl hydrocarbon receptor (AhR) has dual roles in regulating intracellular protein levels both as a ligand-activated transcription factor and as a ligand-dependent E3 ubiquitin ligase. Here, we show that the AhR E3 ubiquitin ligase has a role in suppression of intestinal carcinogenesis by a previously undescribed ligand-dependent beta-catenin degradation pathway that is independent of and parallel to the APC system. This function of AhR is activated by both xenobiotics and natural AhR ligands, such as indole derivatives that are converted from dietary tryptophan and glucosinolates by intestinal microbes, and suppresses intestinal tumor development in Apc(Min/+) mice. These findings suggest that chemoprevention with naturally-occurring and chemically-designed AhR ligands can be used to successfully prevent intestinal cancers.
Proceedings of the National Academy of Sciences 08/2009; 106(32):13481-6. · 9.68 Impact Factor
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Peter Ebbesen,
Erik O Pettersen,
Thomas A Gorr,
Gerhard Jobst,
Kaye Williams,
Jochen Kieninger,
Roland H Wenger,
Silvia Pastorekova,
Ludwig Dubois,
Philippe Lambin, [......],
Twan Van Den Beucken,
Claudiu T Supuran, Lorenz Poellinger,
Peter Ratcliffe,
Arvydas Kanopka,
Agnes Görlach,
Max Gasmann,
Adrian L Harris,
Patrick Maxwell,
Andrea Scozzafava
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ABSTRACT: Cancer cells in hypoxic areas of solid tumors are to a large extent protected against the action of radiation as well as many chemotherapeutic drugs. There are, however, two different aspects of the problem caused by tumor hypoxia when cancer therapy is concerned: One is due to the chemical reactions that molecular oxygen enters into therapeutically targeted cells. This results in a direct chemical protection against therapy by the hypoxic microenvironment, which has little to do with cellular biological regulatory processes. This part of the protective effect of hypoxia has been known for more than half a century and has been studied extensively. However, in recent years there has been more focus on the other aspect of hypoxia, namely the effect of this microenvironmental condition on selecting cells with certain genetic prerequisites that are negative with respect to patient prognosis. There are adaptive mechanisms, where hypoxia induces regulatory cascades in cells resulting in a changed metabolism or changes in extracellular signaling. These processes may lead to changes in cellular intrinsic sensitivity to treatment irrespective of oxygenation and, furthermore, may also have consequences for tissue organization. Thus, the adaptive mechanisms induced by hypoxia itself may have a selective effect on cells, with a fine-tuned protection against damage and stress of many kinds. It therefore could be that the adaptive mechanisms may take advantage of for new tumor labeling/imaging and treatment strategies. One of the Achilles' heels of hypoxia research has always been the exact measurements of tissue oxygenation as well as the control of oxygenation in biological tumor models. Thus, development of technology that can ease this control is vital in order to study mechanisms and perform drug development under relevant conditions. An integrated EU Framework project 2004-2009, termed EUROXY, demonstrates several pathways involved in transcription and translation control of the hypoxic cell phenotype and evidence of cross-talk with responses to pH and redox changes. The carbonic anhydrase isoenzyme CA IX was selected for further studies due to its expression on the surface of many types of hypoxic tumors. The effort has led to marketable culture flasks with sensors and incubation equipment, and the synthesis of new drug candidates against new molecular targets. New labeling/imaging methods for cancer diagnosing and imaging of hypoxic cancer tissue are now being tested in xenograft models and are also in early clinical testing, while new potential anti-cancer drugs are undergoing tests using xenografted tumor cancers. The present article describes the above results in individual consortium partner presentations.
Journal of Enzyme Inhibition and Medicinal Chemistry 05/2009; 24 Suppl 1:1-39. · 1.62 Impact Factor
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ABSTRACT: Tumour-associated expression of CA IX (carbonic anhydrase IX) is to a major extent regulated by HIF-1 (hypoxia-inducible factor-1) which is important for transcriptional activation and consists of the oxygen-regulated subunit HIF-1alpha and the partner factor ARNT [AhR (aryl hydrocarbon receptor) nuclear translocator]. We have previously observed that HIF-1alpha competes with the AhR for interaction with ARNT under conditions when both conditionally regulated factors are activated. We have therefore investigated whether TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin)-induced activation of the AhR pathway might interfere with CA IX expression. The results from the present study suggest that TCDD treatment reduces hypoxic induction of both CA IX mRNA and protein expression. Moreover, the transcriptional activity of the CA9 promoter was significantly reduced by expression of CAAhR (constitutively active AhR), which activates transcription in a ligand-independent manner. Finally, we found that ARNT is critical for both hypoxic induction and the TCDD-mediated inhibition of CA9 expression.
Biochemical Journal 02/2009; 419(2):419-25. · 4.90 Impact Factor
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Ileana Ruxandra Botusan,
Vivekananda Gupta Sunkari,
Octavian Savu,
Anca Irinel Catrina,
Jacob Grünler,
Stina Lindberg,
Teresa Pereira,
Seppo Ylä-Herttuala, Lorenz Poellinger,
Kerstin Brismar,
Sergiu-Bogdan Catrina
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ABSTRACT: Relative hypoxia is essential in wound healing since it normally plays a pivotal role in regulation of all the critical processes involved in tissue repair. Hypoxia-inducible factor (HIF) 1alpha is the critical transcription factor that regulates adaptive responses to hypoxia. HIF-1alpha stability and function is regulated by oxygen-dependent soluble hydroxylases targeting critical proline and asparaginyl residues. Here we show that hyperglycemia complexly affects both HIF-1alpha stability and activation, resulting in suppression of expression of HIF-1 target genes essential for wound healing both in vitro and in vivo. However, by blocking HIF-1alpha hydroxylation through chemical inhibition, it is possible to reverse this negative effect of hyperglycemia and to improve the wound healing process (i.e., granulation, vascularization, epidermal regeneration, and recruitment of endothelial precursors). Local adenovirus-mediated transfer of two stable HIF constructs demonstrated that stabilization of HIF-1alpha is necessary and sufficient for promoting wound healing in a diabetic environment. Our findings outline the necessity to develop specific hydroxylase inhibitors as therapeutic agents for chronic diabetes wounds. In conclusion, we demonstrate that impaired regulation of HIF-1alpha is essential for the development of diabetic wounds, and we provide evidence that stabilization of HIF-1alpha is critical to reverse the pathological process.
Proceedings of the National Academy of Sciences 01/2009; 105(49):19426-31. · 9.68 Impact Factor
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ABSTRACT: Despite a relatively simple core-signaling transduction machinery, Notch signaling controls cell differentiation in many different tissues and at multiple stages in a given cell lineage. To understand how Notch generates this multitude of cellular responses, it is important to learn how the Notch-signaling output is modulated at various levels. Pathway-intrinsic as well as pathway-extrinsic mechanisms, including cross-talk between Notch and other major signaling mechanisms, modulate Notch signaling, contributing to the versatile output. In this review, we discuss how Notch signaling is altered in tumors and illustrate the complexity in signaling pathway cross-talk with examples of how Notch synergizes with NF-kappaB signaling and the cellular response to lowered oxygen (hypoxia).
Current opinion in genetics & development 10/2008; 18(5):449-54. · 8.99 Impact Factor
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Adam T Boutin,
Alexander Weidemann,
Zhenxing Fu,
Lernik Mesropian,
Katarina Gradin,
Colin Jamora,
Michael Wiesener,
Kai-Uwe Eckardt,
Cameron J Koch,
Lesley G Ellies,
Gabriel Haddad,
Volker H Haase,
M Celeste Simon, Lorenz Poellinger,
Frank L Powell,
Randall S Johnson
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ABSTRACT: Skin plays an essential role, mediated in part by its remarkable vascular plasticity, in adaptation to environmental stimuli. Certain vertebrates, such as amphibians, respond to hypoxia in part through the skin; but it is unknown whether this tissue can influence mammalian systemic adaptation to low oxygen levels. We have found that epidermal deletion of the hypoxia-responsive transcription factor HIF-1alpha inhibits renal erythropoietin (EPO) synthesis in response to hypoxia. Conversely, mice with an epidermal deletion of the von Hippel-Lindau (VHL) factor, a negative regulator of HIF, have increased EPO synthesis and polycythemia. We show that nitric oxide release induced by the HIF pathway acts on cutaneous vascular flow to increase systemic erythropoietin expression. These results demonstrate that in mice the skin is a critical mediator of systemic responses to environmental oxygen.
Cell 05/2008; 133(2):223-34. · 32.40 Impact Factor
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ABSTRACT: Tumor hypoxia is linked to increased metastatic potential, but the molecular mechanisms coupling hypoxia to metastasis are poorly understood. Here, we show that Notch signaling is required to convert the hypoxic stimulus into epithelial-mesenchymal transition (EMT), increased motility, and invasiveness. Inhibition of Notch signaling abrogated hypoxia-induced EMT and invasion, and, conversely, an activated form of Notch could substitute for hypoxia to induce these processes. Notch signaling deploys two distinct mechanisms that act in synergy to control the expression of Snail-1, a critical regulator of EMT. First, Notch directly up-regulated Snail-1 expression by recruitment of the Notch intracellular domain to the Snail-1 promoter, and second, Notch potentiated hypoxia-inducible factor 1alpha (HIF-1alpha) recruitment to the lysyl oxidase (LOX) promoter and elevated the hypoxia-induced up-regulation of LOX, which stabilizes the Snail-1 protein. In sum, these data demonstrate a complex integration of the hypoxia and Notch signaling pathways in regulation of EMT and open up perspectives for pharmacological intervention with hypoxiainduced EMT and cell invasiveness in tumors.
Proceedings of the National Academy of Sciences 05/2008; 105(17):6392-7. · 9.68 Impact Factor
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Xiaofeng Zheng,
Sarah Linke,
José M Dias,
Xiaowei Zheng,
Katarina Gradin,
Tristan P Wallis,
Brett R Hamilton,
Maria Gustafsson,
Jorge L Ruas,
Sarah Wilkins,
Rebecca L Bilton,
Kerstin Brismar,
Murray L Whitelaw,
Teresa Pereira,
Jeffrey J Gorman,
Johan Ericson,
Daniel J Peet,
Urban Lendahl, Lorenz Poellinger
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ABSTRACT: Cells adapt to hypoxia by a cellular response, where hypoxia-inducible factor 1alpha (HIF-1alpha) becomes stabilized and directly activates transcription of downstream genes. In addition to this "canonical" response, certain aspects of the pathway require integration with Notch signaling, i.e., HIF-1alpha can interact with the Notch intracellular domain (ICD) to augment the Notch downstream response. In this work, we demonstrate an additional level of complexity in this cross-talk: factor-inhibiting HIF-1 (FIH-1) regulates not only HIF activity, but also the Notch signaling output and, in addition, plays a role in how Notch signaling modulates the hypoxic response. We show that FIH-1 hydroxylates Notch ICD at two residues (N(1945) and N(2012)) that are critical for the function of Notch ICD as a transactivator within cells and during neurogenesis and myogenesis in vivo. FIH-1 negatively regulates Notch activity and accelerates myogenic differentiation. In its modulation of the hypoxic response, Notch ICD enhances recruitment of HIF-1alpha to its target promoters and derepresses HIF-1alpha function. Addition of FIH-1, which has a higher affinity for Notch ICD than for HIF-1alpha, abrogates the derepression, suggesting that Notch ICD sequesters FIH-1 away from HIF-1alpha. In conclusion, the data reveal posttranslational modification of the activated form of the Notch receptor and an intricate mode of cross-coupling between the Notch and hypoxia signaling pathways.
Proceedings of the National Academy of Sciences 04/2008; 105(9):3368-73. · 9.68 Impact Factor
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Toshiharu Yamashita,
Osamu Ohneda,
Masumi Nagano,
Motoyuki Iemitsu,
Yuichi Makino,
Hirotoshi Tanaka,
Takashi Miyauchi,
Katsutoshi Goto,
Kinuko Ohneda,
Yoshiaki Fujii-Kuriyama, Lorenz Poellinger,
Masayuki Yamamoto
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ABSTRACT: Hypoxia-inducible factors (HIFs) are crucial for oxygen homeostasis during both embryonic development and postnatal life. Here we show that a novel HIF family basic helix-loop-helix (bHLH) PAS (Per-Arnt-Sim) protein, which is expressed predominantly during embryonic and neonatal stages and thereby designated NEPAS (neonatal and embryonic PAS), acts as a negative regulator of HIF-mediated gene expression. NEPAS mRNA is derived from the HIF-3alpha gene by alternative splicing, replacing the first exon of HIF-3alpha with that of inhibitory PAS. NEPAS can dimerize with Arnt and exhibits only low levels of transcriptional activity, similar to that of HIF-3alpha. NEPAS suppressed reporter gene expression driven by HIF-1alpha and HIF-2alpha. By generating mice with a targeted disruption of the NEPAS/HIF-3alpha locus, we found that homozygous mutant mice (NEPAS/HIF-3alpha(-)(/)(-)) were viable but displayed enlargement of the right ventricle and impaired lung remodeling. The expression of endothelin 1 and platelet-derived growth factor beta was increased in the lung endothelial cells of NEPAS/HIF-3alpha-null mice. These results demonstrate a novel regulatory mechanism in which the activities of HIF-1alpha and HIF-2alpha are negatively regulated by NEPAS in endothelial cells, which is pertinent to lung and heart development during the embryonic and neonatal stages.
Molecular and cellular biology 03/2008; 28(4):1285-97. · 6.06 Impact Factor
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Steven D Mason,
Helene Rundqvist,
Ioanna Papandreou,
Roger Duh,
Wayne J McNulty,
Richard A Howlett,
I Mark Olfert,
Carl Johan Sundberg,
Nicholas C Denko, Lorenz Poellinger,
Randall S Johnson
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ABSTRACT: During endurance training, exercising skeletal muscle experiences severe and repetitive oxygen stress. The primary transcriptional response factor for acclimation to hypoxic stress is hypoxia-inducible factor-1alpha (HIF-1alpha), which upregulates glycolysis and angiogenesis in response to low levels of tissue oxygenation. To examine the role of HIF-1alpha in endurance training, we have created mice specifically lacking skeletal muscle HIF-1alpha and subjected them to an endurance training protocol. We found that only wild-type mice improve their oxidative capacity, as measured by the respiratory exchange ratio; surprisingly, we found that HIF-1alpha null mice have already upregulated this parameter without training. Furthermore, untrained HIF-1alpha null mice have an increased capillary to fiber ratio and elevated oxidative enzyme activities. These changes correlate with constitutively activated AMP-activated protein kinase in the HIF-1alpha null muscles. Additionally, HIF-1alpha null muscles have decreased expression of pyruvate dehydrogenase kinase I, a HIF-1alpha target that inhibits oxidative metabolism. These data demonstrate that removal of HIF-1alpha causes an adaptive response in skeletal muscle akin to endurance training and provides evidence for the suppression of mitochondrial biogenesis by HIF-1alpha in normal tissue.
AJP Regulatory Integrative and Comparative Physiology 12/2007; 293(5):R2059-69. · 3.34 Impact Factor
