The telomere maintenance enzyme telomerase is a ribonuclear protein complex that is active in 80-90% of immortal cancer cells. Three essential components of the active telomerase holoenzyme include the reverse transcriptase, hTERT, an RNA component (hTR) that has a template region for reverse transcription and the RNA binding and modifying protein, dyskerin. There is evidence indicating that these components may contribute to the survival and/or proliferation of tumour cells via mechanisms that are independent of their roles within the telomerase holoenzyme. The aim of these investigations was to directly compare the functional molecular consequences of abolishing hTERT, hTR and dyskerin in normal and immortal cells in order to determine the most effective and least toxic approach to targeting telomerase.
siRNA was employed to suppress expression of hTERT, hTR and dyskerin in isogenic normal, immortal (pre-malignant) and tumorigenic human foetal lung (MRC5) myofibroblasts, as well as a fibrosarcoma derived cell line, Ht1080. Repression of dyskerin and hTERT caused an immediate halt to the proliferation of the immortal and tumorigenic cells, but had no overt effect on normal cells. Suppression of either dyskerin or hTERT also impaired anchorage-independent growth of tumorigenic cells, while in contrast, treatment with hTR siRNA or a small molecule weight telomerase enzyme inhibitor had no significant effect on any of the cell lines in these short-term assays. Cell cycle analyses indicated that hTERT and dyskerin repression arrested proliferation via distinct mechanisms. Microarray gene expression analysis and Gene Set Enrichment Analyses (GSEA) were employed to unveil divergent mechanisms that underpinned the effects of hTERT and dyskerin repression. Together the results from this study demonstrate that the replication of immortal cells is highly dependent upon continued expression of hTERT and dyskerin, and that these telomerase components support the replication of immortal and tumorigenic cells via distinct mechanisms that are (at least in part) independent of their role in telomere maintenance.