The principal role of Ku in telomere length maintenance is promotion of Est1 association with telomeres.
Bottom Line: The promotion of TLC1 nuclear localization and Est2 recruitment have been proposed to be the principal role of Ku in telomere length maintenance, but neither model has been directly tested.Moreover, restoration of TLC1 nuclear localization, even when combined with Est2 recruitment, does not bypass the role of Ku.Together, our results unexpectedly demonstrate that the principal role of Ku in telomere length maintenance is to promote the association of Est1 with telomeres, which may in turn allow for efficient recruitment and activation of the telomerase holoenzyme.
Affiliation: Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, Texas 77030.Show MeSH
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Mentions: The above results indicate that the recruitment of Est2 to telomeres is not the main role of Ku in telomere elongation. As Est1 recruitment to telomeres in the absence of Ku–TLC1 interaction or Ku is also reduced (Fisher et al. 2004), we next wanted to determine whether expression of a Cdc13–Est1 fusion could bypass the role of Ku. It was previously demonstrated that, after ∼100 generations of growth, telomere lengths in WT and yku70∆ strains expressing a Cdc13–Est1 fusion were similarly elongated (Grandin et al. 2000). These analyses, however, were conducted at high temperature to reveal effects conferred by the cdc13-1 allele also present in the strains, and it is known that telomere and subtelomeric structure is altered in Ku-deficient strains at high temperature (Gravel et al. 1998; Fellerhoff et al. 2000; Maringele and Lydall 2002) due to an apparently naturally thermolabile telomere-specific activity (Paschini et al. 2012). Additionally, the presence of extensive G-tails in the yku70∆ strain would have resulted in a marked increase in binding sites of the Cdc13–Est1 fusion. Therefore, we wanted to test whether defects in Ku–TLC1 interaction alone affects the ability of a Cdc13–Est1 fusion to promote progressive telomere elongation. To do this, we examined telomere length in WT, yku80-135i, and yku80∆ strains expressing a Cdc13–Est1 fusion over successive colony streakouts. In contrast to what was seen in the presence of a Cdc13–Est2 fusion, expression of a Cdc13–Est1 fusion resulted in progressive telomere elongation to approximately the same extent in all strains (Figure 3A and Figure S2). Because a yku80-135i strain does not exhibit extensive G-tails (Stellwagen et al. 2003), this indicates that the extensive elongation observed in the yku80-135i strain was not a consequence of additional Cdc13-binding sites. These results suggest that Ku influences telomere length mainly via Est1.
Affiliation: Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, Texas 77030.