title: Troy, a tumor necrosis factor receptor family member, interacts with lgr5 to inhibit wnt signaling in intestinal stem cells.
abstract: OBJECTIVE
  The Wnt signaling pathway is required for maintenance of the intestinal epithelia; blocking this pathway reduces the proliferative capacity of the intestinal stem cells. However, aberrant Wnt signaling leads to intestinal cancer. We investigated the roles of the Wnt pathway in homeostasis of the intestinal epithelium and during malignant transformation in human cells and mice.
  
  
  METHODS
  We performed chromatin immunoprecipitation (ChIP) with DNA microarray analysis (ChIP-on-chip) to identify genes regulated by Wnt signaling in human colorectal cancer cells Colo320, DLD1, LS174T, and SW480. Formation of intestinal tumor was induced in C57BL/6J mice using azoxymethane and dextran sulfate. Intestinal tissues from these mice, as well as Apc(+/Min) and Apc(CKO/CKO)/Lgr5-EGFP-IRES-CreERT2 mice, were analyzed by immunohistochemistry and in situ hybridization.
  
  
  RESULTS
  We identified promoter regions of 960 genes that interacted with the Wnt pathway nuclear effector T-cell factor 4 in 4 different human colorectal cancer-derived cell lines; 18 of these promoters were present in all chromatin precipitates. Wnt signaling up-regulated a member of the tumor necrosis factor receptor superfamily called TROY. Levels of TROY messenger RNA were increased in human cells with deficiencies in the adenomatous polyposis coli (APC) gene and in cells stimulated with the Wnt3a ligand. Expression of Troy was significantly up-regulated in neoplastic tissues from mice during intestinal tumorigenesis. Lineage tracing experiments revealed that Troy is produced specifically by fast-cycling intestinal stem cells. TROY associated with a unique marker of these cells, leucine-rich repeat-containing G-protein coupled receptor (LGR) 5. In organoids established from the intestinal crypts, Troy suppressed signaling mediated by R-spondin, a Wnt agonist.
  
  
  CONCLUSIONS
  TROY is up-regulated in human colorectal cancer cell lines and in intestinal tumors in mice. It functions as a negative modulator of the Wnt pathway in LGR5-positive stem cells.
authors.affiliation: Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
authors.forename: Bohumil
authors.initials: B
authors.lastname: Fafilek
authors.affiliation: Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
authors.forename: Michaela
authors.initials: M
authors.lastname: Krausova
authors.affiliation: Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
authors.forename: Martina
authors.initials: M
authors.lastname: Vojtechova
authors.affiliation: Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
authors.forename: Vendula
authors.initials: V
authors.lastname: Pospichalova
authors.affiliation: Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
authors.forename: Lucie
authors.initials: L
authors.lastname: Tumova
authors.affiliation: Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
authors.forename: Eva
authors.initials: E
authors.lastname: Sloncova
authors.affiliation: Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
authors.forename: Martina
authors.initials: M
authors.lastname: Huranova
authors.affiliation: Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
authors.forename: Jitka
authors.initials: J
authors.lastname: Stancikova
authors.affiliation: Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
authors.forename: Adela
authors.initials: A
authors.lastname: Hlavata
authors.affiliation: Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic; Second Department of Internal Medicine, Third Faculty of Medicine, Charles University, Prague, Czech Republic.
authors.forename: Jiri
authors.initials: J
authors.lastname: Svec
authors.affiliation: Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
authors.forename: Radislav
authors.initials: R
authors.lastname: Sedlacek
authors.affiliation: Institute for Clinical and Experimental Medicine, Prague, Czech Republic.
authors.forename: Ondrej
authors.initials: O
authors.lastname: Luksan
authors.affiliation: Institute for Clinical and Experimental Medicine, Prague, Czech Republic.
authors.forename: Martin
authors.initials: M
authors.lastname: Oliverius
authors.affiliation: Institute for Clinical and Experimental Medicine, Prague, Czech Republic.
authors.forename: Ludek
authors.initials: L
authors.lastname: Voska
authors.affiliation: Institute for Clinical and Experimental Medicine, Prague, Czech Republic.
authors.forename: Milan
authors.initials: M
authors.lastname: Jirsa
authors.affiliation: Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
authors.forename: Jan
authors.initials: J
authors.lastname: Paces
authors.affiliation: Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
authors.forename: Michal
authors.initials: M
authors.lastname: Kolar
authors.affiliation: Central European Biosystems Ltd, Vestec u Prahy, Czech Republic.
authors.forename: Maria
authors.initials: M
authors.lastname: Krivjanska
authors.affiliation: Institute of Microbiology, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
authors.forename: Klara
authors.initials: K
authors.lastname: Klimesova
authors.affiliation: Institute of Microbiology, Academy of Sciences of the Czech Republic, Prague, Czech Republic.
authors.forename: Helena
authors.initials: H
authors.lastname: Tlaskalova-Hogenova
authors.affiliation: Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, Prague, Czech Republic. Electronic address: korinek@img.cas.cz.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Korinek
mesh_terms: D000818:Animals; D017209:Apoptosis; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002471:Cell Transformation, Neoplastic / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D015179:Colorectal Neoplasms / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D007413:Intestinal Mucosa / Q000378:metabolism* / Q000473:pathology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009374:Neoplasms, Experimental; D014411:Neoplastic Stem Cells / Q000378:metabolism* / Q000473:pathology; D020411:Oligonucleotide Array Sequence Analysis; D012334:RNA, Neoplasm / Q000235:genetics; D043562:Receptors, G-Protein-Coupled / Q000502:physiology*; D018124:Receptors, Tumor Necrosis Factor / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D060449:Wnt Signaling Pathway / Q000502:physiology*
chemical_list: C493597:Lgr5 protein, mouse; D012334:RNA, Neoplasm; D043562:Receptors, G-Protein-Coupled; D018124:Receptors, Tumor Necrosis Factor; C409471:Tnfrsf19 protein, mouse