Telomerase Resets the Homeostatic Telomere Length and Prevents Telomere Dysfunction in Immortalized Human Cells

Abstract

The enzyme telomerase correlates with cellular immortalization, and has been shown to be associated with more than 85% of all human cancers. Introduction of the catalytic subunit of telomerase (hTERT) into normal human cells has been shown to lengthen telomeres and extend life span, while ectopic expression of hTERT in immortal and tumor-derived cells lengthens telomeres even in the presence of endogenous telomerase. Because introduced telomerase has been shown to prevent genomic instability in normal cells, we sought to determine its effects on telomere dynamics and chromosome stability in our immortal cell line. We followed the telomere lengthening of an SV40 T antigen fibroblast cell line (SW39) with overexpressed hTERT for more than a year in culture and found that telomeres continued to elongate before the overall length stabilized. Overexpression of hTERT resets the weighted stable telomere length from an average of 4.5 kb to a final average length of 12.5 kb. The levels of telomerase activity and endogenous expression of hTERT mRNA does not change in these cells over time even after stabilization of telomere length, suggesting that the presence of exogenous hTERT has no effect on endogenous levels. Furthermore, our data suggests a lack of a feedback mechanism for endogenous telomerase expression as a result of hTERT introduction and telomere elongation. The G-rich telomeric overhangs were found to be threefold longer in both the elongating and stabilized populations, while chromosomal end fusion events were reduced by 50% in ectopic hTERT cells. Endogenous expression of the end-protective telomere binding protein, TRF2, increased substantially during telomere elongation and stabilization. Overall, we have established an excellent model to study the role of telomerase and telomere binding proteins in telomere biogenesis by showing that telomerase overexpression resets the homeostatic telomere length and cooperates with TRF2 to prevent genomic instability.