Establishing cell-based reporter systems for the analysis of hTERT expression.

Yi Yuan Huang, Jing Wen Shih, Jing Jer Lin

Research output: Contribution to journalArticle

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Abstract

Telomeres are the protective structures at the end of eukaryotic chromosomes. Telomerase is a ribonucleoprotein that contains both an RNA and a protein component for the maintenance of telomere length. Telomerase activity is detected in the majority of malignant tumors, but not in normal somatic cells, suggesting that telomerase reactivation is a crucial step in cell immortality and carcinogenesis. The mechanism of how telomerase is activated during tumorigenesis remains unclear. However, the expression of the human telomerase reverse transcriptase (hTERT) gene, which encodes the catalytic protein subunit of human telomerase, has been shown to be the major determining factor. To gain insight into the mechanisms regulating hTERT expression and to facilitate the screening of agents that affect hTERT expression, we have established cell-based systems for monitoring hTERT expression. We linked the hTERT promoter to two different reporter genes encoding green fluorescence protein (GFP) and secreted alkaline phosphatase (SEAP), respectively. These constructs were then transfected into H1299 and hTERT-BJ1 cells. Stable clones harboring these DNA constructs were isolated. In these cells, hTERT expression can be monitored through the quantification of GFP or SEAP activity on an automatic plate reader. Using these systems, we have identified several small molecule compounds that affect the expression of telomerase.

Original languageEnglish
Pages (from-to)87-96
Number of pages10
JournalMethods in molecular biology (Clifton, N.J.)
Volume405
Publication statusPublished - Dec 1 2007
Externally publishedYes

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Systems Analysis
Telomerase
Alkaline Phosphatase
Carcinogenesis
Fluorescence
Telomere Homeostasis
Proteins
Ribonucleoproteins
Telomere
Protein Subunits
human TERT protein
Reporter Genes
Catalytic Domain
Clone Cells
Chromosomes
RNA
DNA
Genes
Neoplasms

ASJC Scopus subject areas

  • Genetics
  • Molecular Biology

Cite this

Establishing cell-based reporter systems for the analysis of hTERT expression. / Huang, Yi Yuan; Shih, Jing Wen; Lin, Jing Jer.

In: Methods in molecular biology (Clifton, N.J.), Vol. 405, 01.12.2007, p. 87-96.

Research output: Contribution to journalArticle

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