Identification of the determinants for the specific recognition of single-strand telomeric DNA by Cdc13.
The single-strand overhang present at telomeres plays a critical role in mediating both the capping and telomerase regulation functions of telomeres. The telomere end-binding proteins, Cdc13 in Saccharomyces cerevisiae, Pot1 in higher eukaryotes, and TEBP in the ciliated protozoan Oxytricha nova, exhibit sequence-specific binding to their respective single-strand overhangs. S. cerevisiae telomeres are composed of a heterogeneous mixture of GT-rich telomeric sequence, unlike in higher eukaryotes which have a simple repeat that is maintained with high fidelity. In yeast, the telomeric overhang is recognized by the essential protein Cdc13, which coordinates end-capping and telomerase activities at the telomere. The Cdc13 DNA-binding domain (Cdc13-DBD) binds these telomere sequences with high affinity (3 pM) and sequence specificity. To better understand the basis for this remarkable recognition, we have investigated the binding of the Cdc13-DBD to a series of altered DNA substrates. Although an 11-mer of GT-rich sequence is required for full binding affinity, only three of these 11 bases are recognized with high specificity. This specificity differs from that observed in the other known telomere end-binding proteins, but is well suited to the specific role of Cdc13 at yeast telomeres. These studies expand our understanding of telomere recognition by the Cdc13-DBD and of the unique molecular recognition properties of ssDNA binding.
Eldridge
Aimee M
AM
Department of Chemistry and Biochemistry, UCB 215, University of Colorado, Boulder, Colorado 80309-0215, USA.
Halsey
Wayne A
WA
Wuttke
Deborah S
DS
D001483:Base Sequence; D001665:Binding Sites; D019926:Cyclin B / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000235:genetics* / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D013379:Substrate Specificity; D016615:Telomere / Q000235:genetics*; D013816:Thermodynamics
D019926:Cyclin B; D004277:DNA, Single-Stranded