Abstract
Budding yeast Cdc13-Stn1-Ten1 (CST) complex plays an essential role in telomere protection and maintenance, and has been proposed to be a telomere-specific replication protein A (RPA)-like complex. Previous genetic and structural studies revealed a close resemblance between Stn1-Ten1 and RPA32-RPA14. However, the relationship between Cdc13 and RPA70, the largest subunit of RPA, has remained unclear. Here, we report the crystal structure of the N-terminal OB (oligonucleotide/oligosaccharide binding) fold of Cdc13. Although Cdc13 has an RPA70-like domain organization, the structures of Cdc13 OB folds are significantly different from their counterparts in RPA70, suggesting that they have distinct evolutionary origins. Furthermore, our structural and biochemical analyses revealed unexpected dimerization by the N-terminal OB fold and showed that homodimerization is probably a conserved feature of all Cdc13 proteins. We also uncovered the structural basis of the interaction between the Cdc13 N-terminal OB fold and the catalytic subunit of DNA polymerase α (Pol1), and demonstrated a role for Cdc13 dimerization in Pol1 binding. Analysis of the phenotypes of mutants defective in Cdc13 dimerization and Cdc13-Pol1 interaction revealed multiple mechanisms by which dimerization regulates telomere lengths in vivo. Collectively, our findings provide novel insights into the mechanisms and evolution of Cdc13.
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Acknowledgements
We thank Y Chen and W Deng (University of Michigan, USA) for help at various stages of the project. This work was supported by NIH grants (GM 083015-01 to ML, and GM062631 to NL), an American Cancer Society Research Scholar grant (to ML), NSC 97-2311-B-010-005 from Taiwan National Science Council and NHRI-EX98-9625SI from Taiwan National Health Research Institute (to JL), and Public Service Grant DK074270 (to BCF). ML is a Howard Hughes Medical Institute Early Career Scientist. Use of Life Sciences Collaborative Access Team Sector 21 was supported by the Michigan Economic Development Corporation and the Michigan Technology Tri-Corridor (Grant 085P1000817). Use of the Advanced Photon Source was supported by the US Department of Energy, Office of Science, Office of Basic Energy Sciences, under contract no. DE-AC02-06CH11357.
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Supplementary information
Supplementary information, Figure S1
Alignment of large Cdc13 homologues from Saccharomyces and Kluyveromyces spp. (PDF 8455 kb)
Supplementary information, Figure S2
Structural and biochemical characterization of ScCdc13OB1 dimer. (PDF 133 kb)
Supplementary information, Figure S3
Comparison between monomer Cdc13OB1 (upper panel) and dimer Cdc13OB1 (lower panel) reveals that dimerization of Cdc13OB1 almost completely buries the αB helix at the dimeric interface. (PDF 109 kb)
Supplementary information, Figure S4
In vivo functional analysis of Cdc13 homodimerization. (PDF 108 kb)
Supplementary information, Figure S5
In vivo functional analysis of the Cdc13-Pol1 interaction. (PDF 117 kb)
Supplementary information, Figure S6
The distribution of large and small Cdc13 in the Saccharomycotina subphylum of budding yeast. (PDF 724 kb)
Supplementary information, Table S1
Data collection, phasing and refinement statistics. (PDF 128 kb)
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Sun, J., Yang, Y., Wan, K. et al. Structural bases of dimerization of yeast telomere protein Cdc13 and its interaction with the catalytic subunit of DNA polymerase α. Cell Res 21, 258–274 (2011). https://doi.org/10.1038/cr.2010.138
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DOI: https://doi.org/10.1038/cr.2010.138
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