Telomere dynamics in an immortal human cell line.

Abstract

The integration of transfected plasmid DNA at the telomere of chromosome 13 in an immortalized simian virus 40‐transformed human cell line provided the first opportunity to study polymorphism in the number of telomeric repeat sequences on the end of a single chromosome. Three subclones of this cell line were selected for analysis: one with a long telomere on chromosome 13, one with a short telomere, and one with such extreme polymorphism that no distinct band was discernible. Further subcloning demonstrated that telomere polymorphism resulted from both gradual changes and rapid changes that sometimes involved many kilobases. The gradual changes were due to the shortening of telomeres at a rate similar to that reported for telomeres of somatic cells without telomerase, eventually resulting in the loss of nearly all of the telomere. However, telomeres were not generally lost completely, as shown by the absence of polymorphism in the subtelomeric plasmid sequences. Instead, telomeres that were less than a few hundred base pairs in length showed a rapid, highly heterogeneous increase in size. Rapid changes in telomere length also occurred on longer telomeres. The frequency of this type of change in telomere length varied among the subclones and correlated with chromosome fusion. Therefore, the rapid changes in telomere length appeared occasionally to result in the complete loss of telomeric repeat sequences. Rapid changes in telomere length have been associated with telomere loss and chromosome instability in yeast and could be responsible for the high rate of chromosome fusion observed in many human tumor cell lines.

Repression of an alternative mechanism for lengthening of telomeres in somatic cell hybrids

“…Cleavage of long telomeres to wild type size may occur within a single cell division (Li and Lustig, 1996). Further, tagged telomeres in telomerase-negative immortal human cells were occasionally found to undergo rapid shortening and subsequent rapid relengthening, indicating that telomerase activity is not essential for rapid telomere shortening (Murnane et al, 1994). The telomere loss evidenced in our GM8476HFF5 hybrids suggests that ALT is also not required for rapid shortening as presumably it has already been repressed.…”

“…Some immortalized cell lines, however, do not have any detectable telomerase activity but have a highly characteristic pattern of telomere lengths that range from very short to abnormally long (Murnane et al, 1994;Bryan et al, 1995. This pattern of heterogeneous telomere lengths is maintained over many hundreds of population doublings (Rogan et al, 1995), indicating that telomere maintenance can occur in the absence of detectable telomerase activity.…”

Identification of candidate alternative lengthening of telomeres genes by methionine restriction and RNA interference

“…Most cancers utilize telomerase (Greider and Blackburn, 1985) for this purpose, but a significant minority utilize an alternative lengthening of telomeres (ALT) mechanism (Bryan et al, , 1997. Although details of the molecular mechanism of ALT are largely unknown, previous studies have shown that ALT in human cells involves telomere-telomere recombination (Murnane et al, 1994;Dunham et al, 2000). With a few exceptions (Cerone et al, 2005;Fasching et al, 2005;Marciniak et al, 2005;Brachner et al, 2006), the hallmarks of human ALT cells include (1) a unique pattern of telomere length heterogeneity, with telomeres that range from very short to greater than 50 kb long and (2) the presence of ALT-associated promyelocytic leukemia (PML) nuclear bodies (APBs) containing (TTAGGG)n DNA and telomere-specific binding proteins (Yeager et al, 1999).…”

Telomerase activity during spontaneous immortalization of Li-Fraumeni syndrome skin fibroblasts

“…Telomeres are also important in the maintenance of chromosomal stability (Counter et al, 1992;Murnane et al, 1994). In cells that bypass the normal cellular senescence pathway (Shay et al, 1991b), the reactivation of telomerase occurs when the telomeres have reached a critically shortened length (Harley et al, 1990).…”

“…Currently, there are no reports of telomerasenegative tumor-derived cell lines with short telomeres (58 kb) (Avilion et al, 1996;Harley et al, 1994;Shay and Bacchetti, 1997). However, several examples of experimentally immortalized cultured cell lines have been reported that proliferate inde®nitely with long, heterogenous telomere lengths, without the presence of detectable telomerase activity Duncan et al, 1993;Murnane et al, 1994;Wright et al, 1989;Wright and Shay, 1992).…”