Abstract
BackgroundTelomerase controls telomere homeostasis and cell immortality and is a promising anti-cancer target, but few small molecule telomerase inhibitors have been developed. Reactivated transcription of the catalytic subunit hTERT in cancer cells controls telomerase expression. Better understanding of upstream pathways is critical for effective anti-telomerase therapeutics and may reveal new targets to inhibit hTERT expression.Methodology/Principal FindingsIn a focused promoter screen, several GSK3 inhibitors suppressed hTERT reporter activity. GSK3 inhibition using 6-bromoindirubin-3′-oxime suppressed hTERT expression, telomerase activity and telomere length in several cancer cell lines and growth and hTERT expression in ovarian cancer xenografts. Microarray analysis, network modelling and oligonucleotide binding assays suggested that multiple transcription factors were affected. Extensive remodelling involving Sp1, STAT3, c-Myc, NFκB, and p53 occurred at the endogenous hTERT promoter. RNAi screening of the hTERT promoter revealed multiple kinase genes which affect the hTERT promoter, potentially acting through these factors. Prolonged inhibitor treatments caused dynamic expression both of hTERT and of c-Jun, p53, STAT3, AR and c-Myc.Conclusions/SignificanceOur results indicate that GSK3 activates hTERT expression in cancer cells and contributes to telomere length homeostasis. GSK3 inhibition is a clinical strategy for several chronic diseases. These results imply that it may also be useful in cancer therapy. However, the complex network effects we show here have implications for either setting.
Highlights
Telomerase is a ribonucleoprotein reverse transcriptase which counteracts telomere attrition in dividing cells by synthesising telomere DNA [1]
glycogen synthase kinase 3 (GSK3) activates the hTERT promoter In a focused screen of 79 well characterised kinase inhibitors, A2780 cells were transfected with hTERT reporter construct and 32 h post transfection were exposed to 10 mM each inhibitor for 16 h
The analysis suggests that multiple transcription factors may participate in regulation of hTERT by GSK3, including some or all of Sp1, E2F1, SMAD3, STAT3, HIF-1a, Androgen Receptor (AR), p53, c-Myc, ESR1, AP-1 and NFkB
Summary
Telomerase is a ribonucleoprotein reverse transcriptase which counteracts telomere attrition in dividing cells by synthesising telomere DNA [1]. Telomerase activity requires the catalytic subunit hTERT and the RNA subunit hTERC, which contains the template sequence for reverse transcription. Both gene products are over-expressed in cancer cells relative to somatic cells and in most human cancers. Telomere homeostasis is essential for cell immortalisation and telomerase is an attractive anti-cancer target [2]. Telomerase expression in cancer cells is dependent on aberrant hTERC and hTERT transcription, resulting from multiple events including altered signalling and changes in the promoter chromatin environments relative to normal cells [3]. Telomerase controls telomere homeostasis and cell immortality and is a promising anti-cancer target, but few small molecule telomerase inhibitors have been developed. Better understanding of upstream pathways is critical for effective antitelomerase therapeutics and may reveal new targets to inhibit hTERT expression
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