title: Telomerase variant A279T induces telomere dysfunction and inhibits non-canonical telomerase activity in esophageal carcinomas.
abstract: BACKGROUND
  Although implicated in the pathogenesis of several chronic inflammatory disorders and hematologic malignancies, telomerase mutations have not been thoroughly characterized in human cancers. The present study was performed to examine the frequency and potential clinical relevance of telomerase mutations in esophageal carcinomas.
  
  
  METHODS
  Sequencing techniques were used to evaluate mutational status of telomerase reverse transcriptase (TERT) and telomerase RNA component (TERC) in neoplastic and adjacent normal mucosa from 143 esophageal cancer (EsC) patients. MTS, flow cytometry, time lapse microscopy, and murine xenograft techniques were used to assess proliferation, apoptosis, chemotaxis, and tumorigenicity of EsC cells expressing either wtTERT or TERT variants. Immunoprecipitation, immunoblot, immunofluorescence, promoter-reporter and qRT-PCR techniques were used to evaluate interactions of TERT and several TERT variants with BRG-1 and β-catenin, and to assess expression of cytoskeletal proteins, and cell signaling. Fluorescence in-situ hybridization and spectral karyotyping techniques were used to examine telomere length and chromosomal stability.
  
  
  RESULTS
  Sequencing analysis revealed one deletion involving TERC (TERC del 341-360), and two non-synonymous TERT variants [A279T (2 homozygous, 9 heterozygous); A1062T (4 heterozygous)]. The minor allele frequency of the A279T variant was five-fold higher in EsC patients compared to healthy blood donors (p<0.01). Relative to wtTERT, A279T decreased telomere length, destabilized TERT-BRG-1-β-catenin complex, markedly depleted β-catenin, and down-regulated canonical Wnt signaling in cancer cells; these phenomena coincided with decreased proliferation, depletion of additional cytoskeletal proteins, impaired chemotaxis, increased chemosensitivity, and significantly decreased tumorigenicity of EsC cells. A279T expression significantly increased chromosomal aberrations in mouse embryonic fibroblasts (MEFs) following Zeocin™ exposure, as well as Li Fraumeni fibroblasts in the absence of pharmacologically-induced DNA damage.
  
  
  CONCLUSIONS
  A279T induces telomere dysfunction and inhibits non-canonical telomerase activity in esophageal cancer cells. These findings warrant further analysis of A279T expression in esophageal cancers and premalignant esophageal lesions.
authors.affiliation: Thoracic Surgery Section, Thoracic and GI Oncology Branch; National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: Yuwei
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: National Heart, Lung, and Blood Institute, Bethesda, Maryland, United States of America.
authors.forename: Rodrigo
authors.initials: R
authors.lastname: Calado
authors.affiliation: Thoracic Surgery Section, Thoracic and GI Oncology Branch; National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: Mahadev
authors.initials: M
authors.lastname: Rao
authors.affiliation: Thoracic Surgery Section, Thoracic and GI Oncology Branch; National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: Julie A
authors.initials: JA
authors.lastname: Hong
authors.affiliation: Departments of Pathology and Oncology, Johns Hopkins University of Medicine, Baltimore, Maryland, United States of America.
authors.forename: Alan K
authors.initials: AK
authors.lastname: Meeker
authors.affiliation: National Heart, Lung, and Blood Institute, Bethesda, Maryland, United States of America.
authors.forename: Bogdan
authors.initials: B
authors.lastname: Dumitriu
authors.affiliation: Thoracic Surgery Section, Thoracic and GI Oncology Branch; National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: Scott
authors.initials: S
authors.lastname: Atay
authors.affiliation: Laboratory of Cellular Oncology, National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: Peter J
authors.initials: PJ
authors.lastname: McCormick
authors.affiliation: Laboratory of Experimental Carcinogenesis, National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: Susan H
authors.initials: SH
authors.lastname: Garfield
authors.affiliation: Section of Cancer Genomics, National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: Danny
authors.initials: D
authors.lastname: Wangsa
authors.affiliation: Section of Cancer Genomics, National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: Hesed M
authors.initials: HM
authors.lastname: Padilla-Nash
authors.affiliation: Comparative Molecular Cytogenetics Core Facility, National Cancer Institute, Frederick, Maryland, United States of America.
authors.forename: Sandra
authors.initials: S
authors.lastname: Burkett
authors.affiliation: Thoracic Surgery Section, Thoracic and GI Oncology Branch; National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: Mary
authors.initials: M
authors.lastname: Zhang
authors.affiliation: Thoracic Surgery Section, Thoracic and GI Oncology Branch; National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: Tricia F
authors.initials: TF
authors.lastname: Kunst
authors.affiliation: National Heart, Lung, and Blood Institute, Bethesda, Maryland, United States of America.
authors.forename: Nathan R
authors.initials: NR
authors.lastname: Peterson
authors.affiliation: Thoracic Surgery Section, Thoracic and GI Oncology Branch; National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: Sichuan
authors.initials: S
authors.lastname: Xi
authors.affiliation: Thoracic Surgery Section, Thoracic and GI Oncology Branch; National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: Suzanne
authors.initials: S
authors.lastname: Inchauste
authors.affiliation: Department of Thoracic Surgery, Weill Cornell Medical Center, New York, New York, United States of America.
authors.forename: Nasser K
authors.initials: NK
authors.lastname: Altorki
authors.affiliation: Department of Surgery, University of Saskatchewan, Saskatoon, Saskatchewan, Canada.
authors.forename: Alan G
authors.initials: AG
authors.lastname: Casson
authors.affiliation: Section of Thoracic Surgery, University of Michigan Medical Center, Ann Arbor, Michigan, United States of America.
authors.forename: David G
authors.initials: DG
authors.lastname: Beer
authors.affiliation: Laboratory of Human Carcinogenesis, National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: Curtis C
authors.initials: CC
authors.lastname: Harris
authors.affiliation: Section of Cancer Genomics, National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: Thomas
authors.initials: T
authors.lastname: Ried
authors.affiliation: National Heart, Lung, and Blood Institute, Bethesda, Maryland, United States of America.
authors.forename: Neal S
authors.initials: NS
authors.lastname: Young
authors.affiliation: Thoracic Surgery Section, Thoracic and GI Oncology Branch; National Cancer Institute, Bethesda, Maryland, United States of America.
authors.forename: David S
authors.initials: DS
authors.lastname: Schrump
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D045744:Cell Line, Tumor; D004938:Esophageal Neoplasms / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008297:Male; D051379:Mice; D008819:Mice, Nude; D020125:Mutation, Missense; D009363:Neoplasm Proteins / Q000096:biosynthesis* / Q000235:genetics; D012313:RNA / Q000096:biosynthesis / Q000235:genetics; D012334:RNA, Neoplasm / Q000096:biosynthesis / Q000235:genetics; D019098:Telomerase / Q000096:biosynthesis* / Q000235:genetics; D016615:Telomere / Q000201:enzymology / Q000235:genetics; D059505:Telomere Homeostasis
chemical_list: D009363:Neoplasm Proteins; D012334:RNA, Neoplasm; C422952:telomerase RNA; D012313:RNA; C509186:TERT protein, human; D019098:Telomerase