Reconstitution of telomerase activity in normal human cells leads to elongation of telomeres and extended replicative life span

Abstract

Normal somatic cells have a finite life span [1] and lose telomeric DNA, present at the ends of chromosomes, each time they divide as a function of age in vivo or in culture [2-4]. In contrast, many cancer cells and cell lines established from tumours maintain their telomere length by activation of an RNA-protein complex called telomerase, an enzyme originally discovered in Tetrahymena [5], that synthesizes telomeric repeats [6-8]. These findings have led to the formation of the 'telomere hypothesis', which proposes that critical shortening of telomeric DNA due to the end-replication problem [9] is the signal for the initiation of cellular senescence [10,11]. In yeast, the EST2 gene product, the catalytic subunit of telomerase, is essential for telomere maintenance in vivo [12-14]. The recent cloning of the cDNA encoding the catalytic subunit of human telomerase (hTERT) [15,16] makes it possible to test the telomere hypothesis. In this study, we expressed hTERT in normal human diploid fibroblasts, which lack telomerase activity, to determine whether telomerase activity could be reconstituted leading to extension of replicative life span. Our results show that retroviral-mediated expression of hTERT resulted in functional telomerase activity in normal aging human cells. Moreover, reconstitution of telomerase activity in vivo led to an increase in the length of telomeric DNA and to extension of cellular life span. These findings provide direct evidence in support of the telomere hypothesis, indicating that telomere length is one factor that can determine the replicative life span of human cells.

A Pharmacological Chaperone Molecule Induces Cancer Cell Death by Restoring Tertiary DNA Structures in Mutant hTERT Promoters

“…1 hTERT is the key telomerase holoenzyme subunit that confers telomerase activity. 89,90 Recently it has been shown that specific somatic mutations in the hTERT core promoter element result in a 2-4-fold increase in luciferase activity and that these mutations are commonly found in a number of cancers, most notably in bladder cancer, melanomas, and gliomas, where they are often associated with poor prognosis. [45][46][47] It appears that the level of telomerase activity is the best indicator of patient prognosis.…”

Telomerase reverse transcriptase gene is a direct target of c-Myc but is not functionally equivalent in cellular transformation

“…Growth beyond the replicative senescence checkpoint (Hay¯ick limit) correlates well with genetic lesions that interfere with one or more key cellular mortality pathways, most prominently Myc, Rb and/or p53 Wright and Shay, 1995). Studies addressing the relationship between telomere length, telomerase regulation and replicative capacity have established a critical role for the telomerase catalytic protein component or TERT (for Telomerase Reverse Transcriptase) (Bodnar et al, 1998;Vaziri and Benchimol, 1998;Nakamura et al, 1997;Meyerson et al, 1997;Harrington et al, 1997;Kilian et al, 1997;Nakayama et al, 1998) in immortalization. Speci®cally, normal human somatic cells can acquire the ability to maintain telomeres and replicate well beyond the Hay¯ick limit by stable enforced expression of TERT (Bodnar et al, 1998;Vaziri and Benchimol, 1998).…”

Tissue engineering of blood vessels with endothelial cells differentiated from mouse embryonic stem cells

“…It has been proposed that telomere shortening is the molecular clock that triggers senescence. Previous studies showed that when retinal pigment epithelial cells and foreskin fibroblasts were transfected respectively with the vector encoding the human telomerase catalytic subunit (hTCS), more cells doubling (at least 20 doubling) could be achieved in these transfected cells than non-transfected cells [20,21].Also, the established two cell lines had normal karyotype. In this study, hTERT cDNA was transferred into ES7 cells to give TEC 3 cells and these cells were able to maintain the characteristics of endothelial cells and to proliferate infinitely [14].…”