Delivery of yeast telomerase to a DNA break depends on the recruitment functions of Cdc13 and Est1.
The yeast single-strand TG-repeat telomere binding protein Cdc13 and the telomerase accessory protein Est1 play essential roles in chromosome end replication. To determine whether a proposed Cdc13-Est1 interaction recruits telomerase (Est2), we used a simplified system in which telomere formation was monitored at an HO-induced DNA double-strand break (DSB). Tethering of either Cdc13 or Est1 adjacent to a DSB promoted telomere formation, and tethering of Est1, even in the absence of a DSB, resulted in the recruitment of Est2. Est1 association with a DSB containing an adjacent short TG-repeat sequence depended on the Cdc13-Est1 interaction affected by cdc13-2 and est1-60 mutations, whereas Cdc13 association did not. Similarly, Est2 binding to the DSB also required the Cdc13-Est1 interaction, but not synthesis of new TG repeats at the break site. These data demonstrate a critical role for Est1 in recruiting telomerase to its site of action, in cooperation with the telomere binding protein Cdc13.
Department of Molecular Biology and NCCR "Frontiers in Genetics" Program, University of Geneva, Sciences III, 30, Quai Ernest-Ansermet, CH-1211, Geneva 4, Switzerland.
Alessandro
A
Bianchi
Simona
S
Negrini
David
D
Shore
D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism; D009154:Mutation; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D019098:Telomerase / Q000378:metabolism*; D034501:Telomere-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D015003:Yeasts / Q000235:genetics / Q000378:metabolism
C452219:Cdc13 protein, S cerevisiae; D004268:DNA-Binding Proteins; D029701:Saccharomyces cerevisiae Proteins; D034501:Telomere-Binding Proteins; C081647:high affinity DNA-binding factor, S cerevisiae; D004247:DNA; C099024:EST1 protein, S cerevisiae; C509185:EST2 protein, S cerevisiae; D019098:Telomerase