The telomerase-recruitment domain of the telomere binding protein Cdc13 is regulated by Mec1p/Tel1p-dependent phosphorylation.
The DNA damage-responsive protein kinases ATM and ATR phosphorylate SQ/TQ motifs that lie in clusters in most of their in vivo targets. Budding yeast Cdc13p contains two clusters of SQ/TQ motifs, suggesting that it might be a target of Mec1p/Tel1p (yeast ATR/ATM). Here we demonstrated that the telomerase recruitment domain of Cdc13p is phosphorylated by Mec1p and Tel1p. Gel analysis showed that Cdc13p contains a Mec1/Tel1-dependent post-translational modification. Using an immunoprecipitate (IP)-kinase assay, we showed that Mec1p phosphorylates Cdc13p on serine 225, 249, 255 and 306, and Tel1p phosphorylates Cdc13p on serine 225, 249 and 255 in vitro. Phenotypic analysis in vivo revealed that the mutations in the Cdc13p SQ motifs phosphorylated by Mec1p and Tel1p caused multiple telomere and growth defects. In addition, normal telomere length and growth could be restored by expressing a Cdc13-Est1p hybrid protein. These results demonstrate the telomerase recruitment domain of Cdc13p as an important new telomere-specific target of Mec1p/Tel1p.
Tseng
Shun-Fu
SF
Department of Microbiology, College of Medicine, National Taiwan University, Taipei, Taiwan.
Lin
Jing-Jer
JJ
Teng
Shu-Chun
SC
D004256:DNA Polymerase I / Q000378:metabolism; D047468:Immunoprecipitation; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D009154:Mutation; D010641:Phenotype; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012694:Serine / Q000378:metabolism; D019098:Telomerase / Q000378:metabolism*; D016615:Telomere / Q000737:chemistry; D034501:Telomere-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
C452219:Cdc13 protein, S cerevisiae; D047908:Intracellular Signaling Peptides and Proteins; D029701:Saccharomyces cerevisiae Proteins; D034501:Telomere-Binding Proteins; D012694:Serine; C088937:MEC1 protein, S cerevisiae; D017346:Protein Serine-Threonine Kinases; C513623:TEL1 protein, S cerevisiae; C099024:EST1 protein, S cerevisiae; D019098:Telomerase; D004256:DNA Polymerase I; C083536:POL1 protein, S cerevisiae