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Biology Articles » Cell biology » Concise Review: Telomere Biology in Normal and Leukemic Hematopoietic Stem Cells » The "End-Replication Problem" and Telomerase

The "End-Replication Problem" and Telomerase
- Concise Review: Telomere Biology in Normal and Leukemic Hematopoietic Stem Cells


Semiconservative replication of DNA presents a unique problem:the process only works in the 5' to 3' direction, and DNA polymeraserequires binding of an RNA primer. Olovnikov [14] and Watson[15] predicted the consequences of this long before the telomerewas characterized and termed it the end-replication problem.This anticipated the loss of a small 5' nucleotide segment asDNA synthesis took place, with progressive replication-inducedtelomere shortening (Fig. 1). The cellular consequence of uncheckedtelomere loss is replicative senescence, a viable physiologicalstate but one from which no further cell division can occur.It has been hypothesized that this serves as an evolutionaryconserved tumor-suppressor mechanism, analogous to checkpointsduring cell division [16].

 

The widely conserved enzyme telomerase (via its ability to reinstatelost telomeric DNA-repeat sequence) enables cells to bypassreplicative senescence and can confer cellular immortality [17].The two crucial constituents of telomerase (which together canreconstitute telomerase activity in vitro in a rabbit reticulocyte-lysatemodel [18]) consist of a catalytic protein known as human telomerasereverse transcriptase (hTERT) [19, 20] and an RNA template moleculethat contains a sequence complimentary to the human telomericDNA (hTR) [2123]. Whereas hTR can be detected in mosttissues, hTERT is restricted in its expression [20, 24, 25].Introduction of the latter is sufficient to reconstitute telomeraseactivity, and this has led to the concept that hTERT is theprimary rate-limiting component of telomerase. This is, however,a simplistic view, as hTR can be limiting for telomere maintenancein some circumstances (described below).

Telomerase-deficient murine HSC are considerably less able toundergo serial transplantation than wild-type [26]. Human HSCfrom umbilical cord blood (UCB) (with long telomere sequence)have greater proliferative potential than bone marrow (BM) HSCfrom older donors [27]. Thus, for successful ex vivo expansionof HSC (e.g., for tissue engineering or gene therapy, both long-heldambitions of HSC biologists), the ability to manipulate telomerase(and consequently the telomere) is likely to be critical.

Telomerase activity is highly expressed germline cells, therebyprotecting them against telomere shortening. Human germlinetelomeres are typically maintained at approximately 15 kilobasepairs (kbp) [28]. In contrast, telomerase is not expressed inmost normal somatic cells, which is in line with the observationthat these cells have a finite life span in vitro and undergocellular replicative senescence after 40–80 populationdoublings [29], after considerable telomere shortening. Highlevels of telomerase activity have been detected in transformedcell lines and malignant tumors by the sensitive telomere repeatamplification protocol (TRAP) assay [30]. It has therefore beenpostulated that the process of immortalization typically requiresthe reactivation of telomerase for telomere stabilization andmaintenance in tumor cells [31, 32]. That many leukemias (andindeed other cancers) are thought to arise in a stem cell populationhas given further import to understanding telomere biology andthe role of telomerase in this rare but critically importantcell type.


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