Eirini Trompouki

Publications (11)114.33 Total impact

• Article: Zebrafish globin switching occurs in two developmental stages and is controlled by the LCR.

  Jared J Ganis, Nelson Hsia, Eirini Trompouki, Jill L O de Jong, Anthony DiBiase, Janelle S Lambert, Zhiying Jia, Peter J Sabo, Molly Weaver, Richard Sandstrom, John A Stamatoyannopoulos, Yi Zhou, Leonard I Zon
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  ABSTRACT: Globin gene switching is a complex, highly regulated process allowing expression of distinct globin genes at specific developmental stages. Here, for the first time, we have characterized all of the zebrafish globins based on the completed genomic sequence. Two distinct chromosomal loci, termed major (chromosome 3) and minor (chromosome 12), harbor the globin genes containing α/β pairs in a 5'-3' to 3'-5' orientation. Both these loci share synteny with the mammalian α-globin locus. Zebrafish globin expression was assayed during development and demonstrated two globin switches, similar to human development. A conserved regulatory element, the locus control region (LCR), was revealed by analyzing DNase I hypersensitive sites, H3K4 trimethylation marks and GATA1 binding sites. Surprisingly, the position of these sites with relation to the globin genes is evolutionarily conserved, despite a lack of overall sequence conservation. Motifs within the zebrafish LCR include CACCC, GATA, and NFE2 sites, suggesting functional interactions with known transcription factors but not the same LCR architecture. Functional homology to the mammalian α-LCR MCS-R2 region was confirmed by robust and specific reporter expression in erythrocytes of transgenic zebrafish. Our studies provide a comprehensive characterization of the zebrafish globin loci and clarify the regulation of globin switching.
  Developmental Biology 04/2012; 366(2):185-94. · 4.07 Impact Factor
• Article: Nanog-like regulates endoderm formation through the Mxtx2-Nodal pathway.

  Cong Xu, Zi Peng Fan, Patrick Müller, Rachel Fogley, Anthony DiBiase, Eirini Trompouki, Juli Unternaehrer, Fengzhu Xiong, Ingrid Torregroza, Todd Evans, Sean G Megason, George Q Daley, Alexander F Schier, Richard A Young, Leonard I Zon
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  ABSTRACT: In mammalian embryonic stem cells, the acquisition of pluripotency is dependent on Nanog, but the in vivo analysis of Nanog has been hampered by its requirement for early mouse development. In an effort to examine the role of Nanog in vivo, we identified a zebrafish Nanog ortholog and found that its knockdown impaired endoderm formation. Genome-wide transcription analysis revealed that nanog-like morphants fail to develop the extraembryonic yolk syncytial layer (YSL), which produces Nodal, required for endoderm induction. We examined the genes that were regulated by Nanog-like and identified the homeobox gene mxtx2, which is both necessary and sufficient for YSL induction. Chromatin immunoprecipitation assays and genetic studies indicated that Nanog-like directly activates mxtx2, which, in turn, specifies the YSL lineage by directly activating YSL genes. Our study identifies a Nanog-like-Mxtx2-Nodal pathway and establishes a role for Nanog-like in regulating the formation of the extraembryonic tissue required for endoderm induction.
  Developmental cell 03/2012; 22(3):625-38. · 13.36 Impact Factor
• Article: Linking hematopoietic regeneration to developmental signaling pathways: a story of BMP and Wnt.

  Teresa V Bowman, Eirini Trompouki, Leonard I Zon
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  ABSTRACT: Comment on: Trompouki E, et al. Cell 2011; 147:577-89.
  Cell cycle (Georgetown, Tex.) 02/2012; 11(3):424-5. · 5.36 Impact Factor
• Article: Lineage regulators direct BMP and Wnt pathways to cell-specific programs during differentiation and regeneration.

  Eirini Trompouki, Teresa V Bowman, Lee N Lawton, Zi Peng Fan, Dai-Chen Wu, Anthony DiBiase, Corey S Martin, Jennifer N Cech, Anna K Sessa, Jocelyn L Leblanc, Pulin Li, Ellen M Durand, Christian Mosimann, Garrett C Heffner, George Q Daley, Robert F Paulson, Richard A Young, Leonard I Zon
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  ABSTRACT: BMP and Wnt signaling pathways control essential cellular responses through activation of the transcription factors SMAD (BMP) and TCF (Wnt). Here, we show that regeneration of hematopoietic lineages following acute injury depends on the activation of each of these signaling pathways to induce expression of key blood genes. Both SMAD1 and TCF7L2 co-occupy sites with master regulators adjacent to hematopoietic genes. In addition, both SMAD1 and TCF7L2 follow the binding of the predominant lineage regulator during differentiation from multipotent hematopoietic progenitor cells to erythroid cells. Furthermore, induction of the myeloid lineage regulator C/EBPα in erythroid cells shifts binding of SMAD1 to sites newly occupied by C/EBPα, whereas expression of the erythroid regulator GATA1 directs SMAD1 loss on nonerythroid targets. We conclude that the regenerative response mediated by BMP and Wnt signaling pathways is coupled with the lineage master regulators to control the gene programs defining cellular identity.
  Cell 10/2011; 147(3):577-89. · 32.40 Impact Factor
• Article: Chromatin immunoprecipitation in adult zebrafish red cells.

  Eirini Trompouki, Teresa V Bowman, Anthony Dibiase, Yi Zhou, Leonard I Zon
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  ABSTRACT: Zebrafish has been used for many years as a model to study development and disease. The ability of zebrafish to produce thousand of embryos in a synchronous manner has made zebrafish an invaluable tool for genetic and chemical screens. Since its emergence as an important model organism the molecular tools for studying zebrafish have been limited. In this chapter, we describe a simple method to identify DNA binding sites and chromatin architecture in erythrocytes from adult zebrafish using chromatin immunoprecipitation coupled with next generation sequencing. This technique has been used extensively and successfully in other systems and it will be a useful tool for studying epigenetics in zebrafish.
  Methods in cell biology 01/2011; 104:341-52. · 2.05 Impact Factor
• Article: Thymocyte-specific truncation of the deubiquitinating domain of CYLD impairs positive selection in a NF-kappaB essential modulator-dependent manner.

  Ageliki Tsagaratou, Eirini Trompouki, Sofia Grammenoudi, Dimitris L Kontoyiannis, George Mosialos
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  ABSTRACT: The cylindromatosis tumor suppressor gene (Cyld) encodes a deubiquitinating enzyme (CYLD) with immunoregulatory function. In this study, we evaluated the role of Cyld in T cell ontogeny by generating a mouse (Cyld(Delta9)) with a thymocyte-restricted Cyld mutation that causes a C-terminal truncation of the protein and reciprocates catalytically inactive human mutations. Mutant mice had dramatically reduced single positive thymocytes and a substantial loss of peripheral T cells. The analyses of polyclonal and TCR-restricted thymocyte populations possessing the mutation revealed a significant block in positive selection and an increased occurrence of apoptosis at the double-positive stage. Interestingly, in the context of MHC class I and II restricted TCR transgenes, lack of functional CYLD caused massive deletion of thymocytes that would have been positively selected, which is consistent with an impairment of positive selection. Biochemical analysis revealed that Cyld(Delta9) thymocytes exhibit abnormally elevated basal activity of NF-kappaB and JNK. Most importantly, inactivation of NF-kappaB essential modulator fully restored the NF-kappaB activity of Cyld(Delta9) thymocytes to physiologic levels and rescued their developmental and survival defect. This study identifies a fundamental role for functional CYLD in establishing the proper threshold of activation for thymocyte selection by a mechanism dependent on NF-kappaB essential modulator.
  The Journal of Immunology 08/2010; 185(4):2032-43. · 5.79 Impact Factor
• Article: Small molecule screen in zebrafish and HSC expansion.

  Eirini Trompouki, Leonard I Zon
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  ABSTRACT: The zebrafish (Danio rerio) has emerged as a valuable model organism that is amenable for large-scale chemical and genetic screens. The ability of zebrafish to produce large quantities of synchronized, externally fertilized, transparent embryos makes them ideal for screens, which often are not possible in mammalian models. Signaling pathways important for hematopoiesis are well conserved between zebrafish and mammals, making many targets identified in zebrafish screens applicable to mammals. Hematopoiesis in zebrafish occurs in two waves: the primitive or embryonic wave and the definitive or adult wave. Definitive hematopoietic stem cells arise in the aorta-gonad-mesonephros region (AGM) and express conserved markers such as runx1 and c-myb that allow for the detection of stem cells by whole-mount in situ hybridization (WISH). In this protocol, we will discuss a chemical screen in zebrafish embryos to detect compounds that expand or deplete hematopoietic stem cells (HSCs) in vivo. This type of screen represents a powerful tool to study HSCs in zebrafish.
  Methods in molecular biology (Clifton, N.J.) 01/2010; 636:301-16.
• Article: Truncation of the catalytic domain of the cylindromatosis tumor suppressor impairs lung maturation.

  Eirini Trompouki, Ageliki Tsagaratou, Stylianos K Kosmidis, Pascal Dollé, Jun Qian, Dimitris L Kontoyiannis, Wellington V Cardoso, George Mosialos
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  ABSTRACT: Cyld encodes a 956-amino acid deubiquitinating enzyme (CYLD), which is a negative regulator of nuclear factor kappaB and mitogen-activated protein kinase pathways. Mutations that truncate and inactivate the carboxyl-terminal deubiquitinating domain of CYLD underlie the development of skin appendage tumors in humans, whereas down-regulation of Cyld expression has been associated with the development of various types of human malignancies including lung cancer. To establish an animal model of human CYLD inactivation and characterize the biological role of CYLD in vivo, we generated mice carrying a homozygous deletion of Cyld exon 9 (Cyld(Delta 9/Delta 9) mice) using a conditional approach. Deletion of exon 9 would cause a carboxyl-terminal truncation of CYLD and inactivation of its deubiquitinating activity. In accordance with previous studies, fibroblasts from Cyld(Delta 9/Delta 9) embryos had hyperactive nuclear factor kappaB and c-Jun kinase pathways compared with control fibroblasts. Cyld(Delta 9/Delta 9) newborn mice were smaller than wild-type littermates with a short and kinky tail and no major developmental defects. However, Cyld(Delta 9/Delta 9) mice died shortly after birth from apparent respiratory dysfunction. Histological examination of E18.5 Cyld(Delta 9/Delta 9) lungs demonstrated an immature phenotype characterized by hyperplasic mesenchyme but apparently normal epithelial, smooth muscle. and endothelial structures. Our study identifies an important role of CYLD in lung maturation, which may underlie the development of many cases of lung cancer.
  Neoplasia (New York, N.Y.) 06/2009; 11(5):469-76. · 5.48 Impact Factor
• Article: NF-kappaB is essential for induction of CYLD, the negative regulator of NF-kappaB: evidence for a novel inducible autoregulatory feedback pathway.

  Hirofumi Jono, Jae Hyang Lim, Lin-Feng Chen, Haidong Xu, Eirini Trompouki, Zhixing K Pan, George Mosialos, Jian-Dong Li
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  ABSTRACT: The transcription factor NF-kappaB regulates genes involved in inflammatory and immune responses, tumorigenesis, and apoptosis. In contrast to the pleiotropic stimuli that lead to its positive regulation, the known signaling mechanisms that underlie the negative regulation of NF-kappaB are very few. Recent studies have identified the tumor suppressor CYLD, loss of which causes a benign human syndrome called cylindromatosis, as a key negative regulator for NF-kappaB signaling by deubiquitinating tumor necrosis factor (TNF) receptor-associated factor (TRAF) 2, TRAF6, and NEMO (NF-kappaB essential modulator, also known as IkappaB kinase gamma). However, how CYLD is regulated remains unknown. The present study revealed a novel autoregulatory feedback pathway through which activation of NF-kappaB by TNF-alpha and bacterium nontypeable Haemophilus influenzae (NTHi) induces CYLD that in turn leads to the negative regulation of NF-kappaB signaling. In addition, TRAF2 and TRAF6 appear to be differentially involved in NF-kappaB-dependent induction of CYLD by TNF-alpha and NTHi. These findings provide novel insights into the autoregulation of NF-kappaB activation.
  Journal of Biological Chemistry 09/2004; 279(35):36171-4. · 4.77 Impact Factor
• Article: NF-κB Is Essential for Induction of CYLD, the Negative Regulator of NF-κB

  Hirofumi Jono, Jae Hyang Lim, Lin-Feng Chen, Haidong Xu, Eirini Trompouki, Zhixing K. Pan, George Mosialos, Jian-Dong Li
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  ABSTRACT: The transcription factor NF-κB regulates genes involved in inflammatory and immune responses, tumorigenesis, and apoptosis. In contrast to the pleiotropic stimuli that lead to its positive regulation, the known signaling mechanisms that underlie the negative regulation of NF-κB are very few. Recent studies have identified the tumor suppressor CYLD, loss of which causes a benign human syndrome called cylindromatosis, as a key negative regulator for NF-κB signaling by deubiquitinating tumor necrosis factor (TNF) receptor-associated factor (TRAF) 2, TRAF6, and NEMO (NF-κB essential modulator, also known as IκB kinase γ). However, how CYLD is regulated remains unknown. The present study revealed a novel autoregulatory feedback pathway through which activation of NF-κB by TNF-α and bacterium nontypeable Haemophilus influenzae (NTHi) induces CYLD that in turn leads to the negative regulation of NF-κB signaling. In addition, TRAF2 and TRAF6 appear to be differentially involved in NF-κB-dependent induction of CYLD by TNF-α and NTHi. These findings provide novel insights into the autoregulation of NF-κB activation.
  Journal of Biological Chemistry 08/2004; 279(35):36171-36174. · 4.77 Impact Factor
• Article: CYLD is a deubiquitinating enzyme that negatively regulates NF-kappaB activation by TNFR family members.

  Eirini Trompouki, Eudoxia Hatzivassiliou, Theodore Tsichritzis, Hannah Farmer, Alan Ashworth, George Mosialos
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  ABSTRACT: Familial cylindromatosis is an autosomal dominant predisposition to tumours of skin appendages called cylindromas. Familial cylindromatosis is caused by mutations in a gene encoding the CYLD protein of previously unknown function. Here we show that CYLD is a deubiquitinating enzyme that negatively regulates activation of the transcription factor NF-kappaB by specific tumour-necrosis factor receptors (TNFRs). Loss of the deubiquitinating activity of CYLD correlates with tumorigenesis. CYLD inhibits activation of NF-kappaB by the TNFR family members CD40, XEDAR and EDAR in a manner that depends on the deubiquitinating activity of CYLD. Downregulation of CYLD by RNA-mediated interference augments both basal and CD40-mediated activation of NF-kappaB. The inhibition of NF-kappaB activation by CYLD is mediated, at least in part, by the deubiquitination and inactivation of TNFR-associated factor 2 (TRAF2) and, to a lesser extent, TRAF6. These results indicate that CYLD is a negative regulator of the cytokine-mediated activation of NF-kappaB that is required for appropriate cellular homeostasis of skin appendages.
  Nature 09/2003; 424(6950):793-6. · 36.28 Impact Factor