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Sensitivity of yeast strains with long G-tails to levels of telomere-bound telomerase.

Authors:
Vega LR, Phillips JA, Thornton BR, Benanti JA, Onigbanjo MT, Toczyski DP, Zakian VA
Affiliation:
Journal:
PLoS genetics

Abstract

The Saccharomyces cerevisiae Pif1p helicase is a negative regulator of telomere length that acts by removing telomerase from chromosome ends. The catalytic subunit of yeast telomerase, Est2p, is telomere associated throughout most of the cell cycle, with peaks of association in both G1 phase (when telomerase is not active) and late S/G2 phase (when telomerase is active). The G1 association of Est2p requires a specific interaction between Ku and telomerase RNA. In mutants lacking this interaction, telomeres were longer in the absence of Pif1p than in the presence of wild-type PIF1, indicating that endogenous Pif1p inhibits the active S/G2 form of telomerase. Pif1p abundance was cell cycle regulated, low in G1 and early S phase and peaking late in the cell cycle. Low Pif1p abundance in G1 phase was anaphase-promoting complex dependent. Thus, endogenous Pif1p is unlikely to act on G1 bound Est2p. Overexpression of Pif1p from a non-cell cycle-regulated promoter dramatically reduced viability in five strains with impaired end protection (cdc13-1, yku80Delta, yku70Delta, yku80-1, and yku80-4), all of which have longer single-strand G-tails than wild-type cells. This reduced viability was suppressed by deleting the EXO1 gene, which encodes a nuclease that acts at compromised telomeres, suggesting that the removal of telomerase by Pif1p exposed telomeres to further C-strand degradation. Consistent with this interpretation, depletion of Pif1p, which increases the amount of telomere-bound telomerase, suppressed the temperature sensitivity of yku70Delta and cdc13-1 cells. Furthermore, eliminating the pathway that recruits Est2p to telomeres in G1 phase in a cdc13-1 strain also reduced viability. These data suggest that wild-type levels of telomere-bound telomerase are critical for the viability of strains whose telomeres are already susceptible to degradation.

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