期刊
G3-GENES GENOMES GENETICS
卷 11, 期 12, 页码 -出版社
OXFORD UNIV PRESS INC
DOI: 10.1093/g3journal/jkab359
关键词
telomerase; end-joining; recombination; replication; checkpoint; DNA repair; senescence
资金
- National Institutes of Health [1R15GM139093-01]
- NIGMS funding from the South Texas Doctoral Bridge Program [R25-GM-102783]
In this study, it was found that overexpression of the Est2 reverse transcriptase and TLC1 RNA template subunits of telomerase, but not other telomerase- and telomere-associated proteins, restored viability of yku mutant cells. Additionally, three catalytically inactive reverse transcriptase mutants were able to efficiently suppress cell death in yku cells. These findings suggest a replication-independent mechanism involving structural reinforcement of protective telomere cap structures in stabilizing chromosomes in yku mutants.
The Ku complex performs multiple functions inside eukaryotic cells, including protection of chromosomal DNA ends from degradation and fusion events, recruitment of telomerase, and repair of double-strand breaks (DSBs). Inactivation of Ku complex genes YKU70 or YKU80 in cells of the yeast Saccharomyces cerevisiae gives rise to mutants that exhibit shortened telomeres and temperature-sensitive growth. In this study, we have investigated the mechanism by which overexpression of telomerase suppresses the temperature sensitivity of yku mutants. Viability of yku cells was restored by overexpression of the Est2 reverse transcriptase and TLC1 RNA template subunits of telomerase, but not the Est1 or Est3 proteins. Overexpression of other telomerase- and telomere-associated proteins (Cdc13, Stn1, Ten1, Rif1, Rif2, Sir3, and Sir4) did not suppress the growth defects of yku70 cells. Mechanistic features of suppression were assessed using several TLC1 RNA deletion derivatives and Est2 enzyme mutants. Supraphysiological levels of three catalytically inactive reverse transcriptase mutants (Est2-D530A, Est2-D670A, and Est2-D671A) suppressed the loss of viability as efficiently as the wild-type Est2 protein, without inducing cell senescence. Roles of proteins regulating telomere length were also determined. The results support a model in which chromosomes in yku mutants are stabilized via a replication-independent mechanism involving structural reinforcement of protective telomere cap structures.
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