Interactions of TLC1 (Which Encodes the RNA Subunit of Telomerase), TEL1, and MEC1 in Regulating Telomere Length in the Yeast Saccharomyces cerevisiae

Abstract

In the yeast Saccharomyces cerevisiae, chromosomes terminate with a repetitive sequence [poly(TG 1-3 )] 350 to 500 bp in length. Strains with a mutation of TEL1, a homolog of the human gene (ATM) mutated in patients with ataxia telangiectasia, have short but stable telomeric repeats. Mutations of TLC1 (encoding the RNA subunit of telomerase) result in strains that have continually shortening telomeres and a gradual loss of cell viability; survivors of senescence arise as a consequence of a Rad52p-dependent recombination events that amplify telomeric and subtelomeric repeats. We show that a mutation in MEC1 (a gene related in sequence to TEL1 and ATM) reduces telomere length and that tel1 mec1 double mutant strains have a senescent phenotype similar to that found in tlc1 strains. As observed in tlc1 strains, survivors of senescence in the tel1 mec1 strains occur by a Rad52p-dependent amplification of telomeric and subtelomeric repeats. In addition, we find that strains with both tel1 and tlc1 mutations have a delayed loss of cell viability compared to strains with the single tlc1 mutation. This result argues that the role of Tel1p in telomere maintenance is not solely a direct activation of telomerase.Most eukaryotic chromosomes end with simple repetitive DNA sequences (7). In the yeast Saccharomyces cerevisiae, wild-type strains have poly(TG 1-3 ) tracts 350 to 500 bp in length (31,42,43). Within a yeast cell population, telomeric tracts, even for a single chromosome, range in size by about Ϯ 50 bp (43). This variability in telomere length, as well as the identification of mutants with telomeric tracts that are longer or shorter than those in wild-type strains, suggests that telomere length is likely to reflect a balance between mechanisms that extend or contract these terminal repeats.In many eukaryotes, including yeast, the most important mechanism for extending telomeric repeats is telomerase (7,45). This RNA-protein enzyme complex extends the G-rich strand of the telomere 5Ј to 3Ј, using telomeric repeats encoded within the RNA component of the enzyme as a template. The extended G-rich strand is thought to be copied 5Ј to 3Ј by conventional DNA polymerases to yield the complementary C-rich strand (7). In yeast, the RNA component of telomerase is encoded by TLC1 (33), and the protein component with reverse transcriptase activity is encoded by EST2 (16). In strains with mutations in either of these genes, telomeric tracts shorten with each cell division, resulting in gradual loss of cell viability (13,33). This same senescent phenotype is observed in strains with mutations in several other EST (ever shorter telomeres) genes, including EST1, EST3, and CDC13/EST4 (13,19). Since mutations in all EST genes have the same phenotype and since double-mutant est strains have the same phenotype as the single mutants, it has been suggested that all Est proteins function in the same pathway of telomere maintenance (13, 23, 40), although not necessarily as components of telomerase (15).Although yeast strains with a mutatio...

The YeastTEL1Gene Partially Substitutes for HumanATMin Suppressing Hyperrecombination, Radiation-Induced Apoptosis and Telomere Shortening in A-T Cells

“…The short telomere phenotype is found in both A-T cells that express strong telomerase activity (e.g., the SV40-transformed fibroblasts used for our experiments) and in A-T cells that lack telomerase activity (primary diploid fibroblasts and lymphocytes) (e.g., Xia et al, 1996). This conclusion is further supported by recent findings in S. cerevisiae that indicate that the role of Tel1p in telomere length control is at least partially independent of telomerase function (Ritchie et al, 1999). The telomere lengthening seen in TEL1-transfected A-T cells is also unlikely to be the result of activating the so-called ALT pathway, in that our stably TEL1-transfected A-T cells did not show the extensive heterogeneity and bimodal distribution of telomere length that is characteristic for this mechanism of telomere extension (Murnane et al, 1994;Bryan et al, 1995;Sprung et al, 1997).…”

“…Additional support for a functional role of ATM in controlling telomere length is provided by the demonstration that expression of a dominant-negative ATM fragment in normal cells results in a decrease in average telomere repeat length (Smilenov et al, 1997). In yeast, deficiency of the homologous TEL1 gene also leads to gradual telomere shortening (Lustig and Petes, 1986;Greenwell et al, 1995;Ritchie et al, 1999).…”

Cdc13 Telomere Capping Decreases Mec1 Association but Does Not Affect Tel1 Association with DNA Ends

“…The Cdc13 telomere cap blocks MRX-dependent DNA degradation and in turn decreases Mec1 accumulation at the DNA end, whereas it permits the MRX complex to recruit Tel1 to the DNA end. Although Tel1 plays a minor role in checkpoint response (Morrow et al, 1995;Sanchez et al, 1996), it plays a predominant role in telomere length regulation (Greenwell et al, 1995;Ritchie et al, 1999). Consistently, telomere synthesis from TG 81 ends largely depends on Tel1 (Frank et al, 2006).…”

“…The maintenance of telomere structure and length is crucial for telomere homeostasis, because dysfunctional or shortened telomeres activate a checkpoint response that is mediated through the ATM and ATR family proteins (d'Adda di Fagagna et al, 2004;de Lange, 2005). However, the ATM and ATR family proteins are also required for proper maintenance of telomeres (Greenwell et al, 1995;Metcalfe et al, 1996;Ritchie et al, 1999). Recent evidence suggests that addition of telomeric TG repeats adjacent to DSBs inhibits the activation of checkpoint response in budding yeast (Michelson et al, 2005).…”

Pol12, the B subunit of DNA polymerase α, functions in both telomere capping and length regulation

“…Previous studies have shown that deletion of either TEL1 (the yeast ATM homolog) or the MRX complex genes RAD50 and MRE11 causes severe telomere shortening (Greenwell et al 1995;Boulton and Jackson 1998), and at least in the case of tel1⌬ a loss of Rap1 counting (Ray and Runge 1999a;Brevet et al 2003). Both Tel1 and MRX are thus thought to be positive regulators of telomerase action, and epistasis analysis would suggest that they act downstream from both RIF1 and RIF2 (Ray and Runge 1999a;Ritchie et al 1999;Ritchie and Petes 2000;Chan et al 2001;Diede and Gottschling 2001;Tsukamoto et al 2001). Significantly, the pol12-216 mutation causes telomere elongation in all three mutant backgrounds (tel1⌬, mre11⌬, and rad50⌬), to slightly longer than wild-type lengths (Fig.…”