Rapid upregulation of telomerase activity in human leukemia HL-60 cells treated with clinical doses of the DNA-damaging drug etoposide

Abstract

The enzyme telomerase is implicated in cellular resistance to apoptosis, but the mechanism for this resistance remains to be elucidated. The ability of telomerase to synthesize new DNA at telomeres suggests that this enzyme might function in the repair of double-stranded DNA breaks. To distinguish the effects of double-stranded DNA break damage and apoptosis on human telomerase activity, we treated the HL-60 human hematopoietic cancer cell line with clinical doses of the chemotherapeutic drug etoposide (0.5 to 5 M), which allowed us to distinguish between events associated with DNA damageinduced cell cycle arrest, and events associated with apoptosis. Large (three-to seven-fold) upregulation of telomerase activity occurred soon after etoposide treatment (3 h) in S/G2/M-arresting populations; this upregulation was abolished at onset of apoptotic cell death. No upregulation of telomerase activity was observed in cells treated with a larger dose of etoposide (5 M) that caused cells to undergo rapid apoptosis without intervening cell cycle arrests. These observations are consistent with a possible role for telomerase upregulation during the DNA damage response.

DNA damage transiently increases TRF2 mRNA expression and telomerase activity

“…HTERT protein levels were reported to be unchanged. 28 Conflicting results were reported by another group who observed downregulation of telomerase activity by etoposide treatment that occurred despite increased hTERT expression in Jurkat, Raji and CEM-6 cells. 30 To gain insights into the effect of etoposide on telomerase regulation, we treated HL60 cells with various concentrations of etoposide and measured telomerase activity, telomere binding protein expression, DNA damage and cell cycle phase distribution.…”

“…Interestingly, overexpression of functional TRF2 can overcome the senescence-inducing signals from critically short telomeres and thus allow cell growth in the presence of short and therefore 'damaged' telomeres. 27 Telomerase activity can be induced by treating cell lines with etoposide, as it has been shown by one group of investigators for HL60 cells 28 and by another for five pancreatic cell lines. 29 The induction of activity was abolished with the onset of apoptosis, and the early time at which telomerase was upregulated suggested an association with DNA damage rather than cell cycle arrest, although direct evidence was lacking.…”

“…In agreement with the results of other authors, no association between high telomerase activity and the G2/M phase was detected. 28 …”

“…28 To further characterize changes at the telomere associated with etoposide treatment, we quantified the expression levels of the best-characterized telomere binding proteins, TRF1 and TRF2, after etoposide treatment by real-time RT-PCR. We found significant upregulation of TRF2 mRNA expression immediately after etoposide treatment, whereas TRF1 mRNA levels remained unchanged.…”

“…29 The induction of activity was abolished with the onset of apoptosis, and the early time at which telomerase was upregulated suggested an association with DNA damage rather than cell cycle arrest, although direct evidence was lacking. 28 Detection of hTERT mRNA or telomere binding protein mRNA levels and direct measurement of DNA damage, for example by the comet assay, were not performed. HTERT protein levels were reported to be unchanged.…”