Schizosaccharomyces pombe Ccq1 and TER1 bind the 14-3-3-like domain of Est1, which promotes and stabilizes telomerase-telomere association.
The telomerase protein Est1 exists in multiple organisms, including Schizosaccharomyces pombe, humans, and Saccharomyces cerevisiae, but its function has only been closely examined in S. cerevisiae, where it is a recruiter/activator of telomerase. Here, we demonstrate that S. pombe Est1 was required for the telomere association of the telomerase holoenzyme, suggesting that it too has a recruitment role. Its association with telomeres was dependent on Trt1, the catalytic subunit, and Ccq1, a telomeric protein. Surprisingly, Est1 telomere binding was only partially dependent on TER1, the telomerase RNA, even though Est1 bound nucleotides 415-507 of TER1. A ter1-Δ415-507 strain had short telomeres and very low Est1 and Trt1 telomere association in late S phase but did not senesce. An unbiased search for mutations that reduced Est1-TER1 interaction identified mutations only in the Est1 14-3-3-like domain, a phosphoserine-binding motif, the first example of a 14-3-3-like domain with RNA-binding activity. These mutations also reduced Est1-Ccq1 binding. One such mutant prevented Est1 telomere association and caused telomere loss and slow senescence, similar to ccq1Δ. We propose that the Est1-Ccq1 interaction is critical for telomerase recruitment, while the Est1-TER1 interaction acts downstream from Ccq1-mediated recruitment to stabilize the holoenzyme at the telomere.
Department of Molecular Biology, Princeton University, Princeton, New Jersey 08544, USA.
Christopher J
CJ
Webb
Virginia A
VA
Zakian
D048948:14-3-3 Proteins / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D011485:Protein Binding; D055550:Protein Stability; D017434:Protein Structure, Tertiary; D012313:RNA; D012568:Schizosaccharomyces / Q000201:enzymology / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019098:Telomerase / Q000378:metabolism*; D016615:Telomere / Q000378:metabolism*; D034501:Telomere-Binding Proteins / Q000378:metabolism*
D048948:14-3-3 Proteins; C529647:Ccq1 protein, S pombe; D004268:DNA-Binding Proteins; D029702:Schizosaccharomyces pombe Proteins; D034501:Telomere-Binding Proteins; C516544:Ter1 telomerase subunit, S pombe; D012313:RNA; C569590:EST1 protein, S pombe; D019098:Telomerase