MEC3,MEC1,andDDC2Are Essential Components of a Telomere Checkpoint Pathway Required for Cell Cycle Arrest during Senescence inSaccharomyces cerevisiae

Abstract

When telomerase is absent and/or telomeres become critically short, cells undergo a progressive decline in viability termed senescence. The telomere checkpoint model predicts that cells will respond to a damaged or critically short telomere by transiently arresting and activating repair of the telomere. We examined the senescence of telomerase-deficient Saccharomyces cerevisiae at the cellular level to ask if the loss of telomerase activity triggers a checkpoint response. As telomerasedeficient mutants were serially subcultured, cells exhibited a progressive decline in average growth rate and an increase in the number of cells delayed in the G2/M stage of the cell cycle. MEC3, MEC1, and DDC2, genes important for the DNA damage checkpoint response, were required for the cell cycle delay in telomerase-deficient cells. In contrast, TEL1, RAD9, and RAD53, genes also required for the DNA damage checkpoint response, were not required for the G2/M delay in telomerase-deficient cells. We propose that the telomere checkpoint is distinct from the DNA damage checkpoint and requires a specific set of gene products to delay the cell cycle and presumably to activate telomerase and/or other telomere repair activities. INTRODUCTIONTelomeres, the nucleotide-protein structures at the ends of linear chromosomes, serve as a cap to protect the ends of chromosomes (reviewed in Blackburn, 2000). The function of this cap must strike a balance between facilitating and limiting access to the telomere. The cap must allow access to telomeric DNA for DNA replication enzymes, replication forks, and telomerase. In contrast, access to other factors that degrade and/or modify DNA ends must be limited. Thus, the cap must be dynamic, coordinating access to telomere DNA with other cellular events such as DNA replication or mitosis.Telomere DNA is normally replicated by telomerase, a specialized reverse transcriptase that utilizes an RNA template that is an integral component of the enzyme. Telomerase is activated late in S phase, around the time when telomeres are replicated (Wellinger et al., 1993a(Wellinger et al., , 1993b. Cells that are telomerase-deficient due to mutations in the catalytic component of the enzyme, the template RNA, or other required factors undergo senescence, a progressive loss of viability that is dependent on the number of divisions after loss of telomerase (Lundblad and Szostak, 1989;McEachern and Blackburn, 1996;Nakamura et al., 1997). During senescence, telomeres become progressively shorter as population viability declines (Singer and Gottschling, 1994;Lendvay et al., 1996;McEachern and Blackburn, 1996;Lingner et al., 1997). In yeasts, senescence can be detected as reduced numbers of colony-forming units and decreased colony size on solid media or as a decline in the average culture growth rate in liquid cultures (Singer and Gottschling, 1994;Lendvay et al., 1996;McEachern and Blackburn, 1996;.Changes in telomere structure can cause cellular abnormalities. In human cells, overexpression of a dominantnegative form of TR...

ELG1 , a regulator of genome stability, has a role in telomere length regulation and in silencing

“…We have also found that the increase in telomeric silencing seen in the absence of ELG1 depended on yKu70p, which is known to recruit the Sir complex to the telomeres (15,39), and on Sir2p, a major component of the telomeric silencing complex (40). Elg1p probably affects silencing directly because we have shown that the telomere position effect enhancement in ⌬elg1 strains is not a consequence of telomere elongation.…”

Activation of a LTR-retrotransposon by telomere erosion

“…This nucleoprotein complex protects the ends of chromosomes from being recognized as double-strand breaks, thereby preventing recombination between telomeres that leads to chromosomal fusions and instability (11,12). In the absence of telomerase, telomeres shorten with every round of DNA replication, eventually reaching a critically short length that triggers arrest in the G 2 ͞M stage of the cell cycle (13)(14)(15)(16). Although the majority of cells cease dividing after 50-100 generations, a fraction of cells escape arrest by forming alternative telomere structures containing either tandem arrays of the subtelomeric repeat, YЈ (type I survivors), or long and heterogeneous tracts of telomeric repeat DNA (type II survivors) (14,17,18).…”

“…The signaling pathway that triggers G 2 ͞M arrest in response to telomere shortening has recently been defined (15,16). The ''telomere checkpoint pathway,'' which is a segment of the DNA-damage checkpoint pathway (reviewed in refs.…”

“…A second arm of the DNA-damage checkpoint pathway that includes Mre11, Rad50, Xrs2, and the Tel1 kinase as sensors is not required for the telomere checkpoint. Interestingly, DNA-damage transducer proteins Rad9 and Rad53 are also dispensable for the telomere checkpoint; however, they are activated in response to telomere erosion and required for induction of a DNA damage-response gene (15,16).…”

Fungal physiology and the origins of molecular biology

“…Many decades later, telomeres were identified as the structures at the ends of chromosomes whose function is to stabilize these chromosomes and prevent stepwise loss of genetic material as a result of the incomplete replication of DNA strands with each round of replication. The discovery of the ribonucleoprotein telomerase, with its enzyme-associated RNA template, which recognizes and extends the G-rich strand of the terminal repeat by adding T-G polydeoxyribonucleotide repeats, came from studies of the ciliate Tetrahymena, but this insight was soon expanded by studies of telomeres and the proteins involved in telomere function in S. cerevisiae by laboratories of V. A. Zakian, B. Tye, Judith Berman, J. E. Haber, E. H. Blackburn, D. Shore, and many others (Enomoto et al, 2002).…”