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The OB-fold domain 1 of human POT1 recognizes both telomeric and non-telomeric DNA motifs.

Biochimie (2015-05-03)
Kyung H Choi, Amanda S Lakamp-Hawley, Carol Kolar, Ying Yan, Gloria E O Borgstahl, Michel M Ouellette
ABSTRACT

The POT1 protein plays a critical role in telomere protection and telomerase regulation. POT1 binds single-stranded 5'-TTAGGGTTAG-3' and forms a dimer with the TPP1 protein. The dimer is recruited to telomeres, either directly or as part of the Shelterin complex. Human POT1 contains two Oligonucleotide/Oligosaccharide Binding (OB) fold domains, OB1 and OB2, which make physical contact with the DNA. OB1 recognizes 5'-TTAGGG whereas OB2 binds to the downstream TTAG-3'. Studies of POT1 proteins from other species have shown that some of these proteins are able to recognize a broader variety of DNA ligands than expected. To explore this possibility in humans, we have used SELEX to reexamine the sequence-specificity of the protein. Using human POT1 as a selection matrix, high-affinity DNA ligands were selected from a pool of randomized single-stranded oligonucleotides. After six successive rounds of selection, two classes of high-affinity targets were obtained. The first class was composed of oligonucleotides containing a cognate POT1 binding sites (5'-TTAGGGTTAG-3'). The second and more abundant class was made of molecules that carried a novel non-telomeric consensus: 5'-TNCANNAGKKKTTAGG-3' (where K = G/T and N = any base). Binding studies showed that these non-telomeric sites were made of an OB1-binding motif (TTAGG) and a non-telomeric motif (NT motif), with the two motifs recognized by distinct regions of the OB1 domain. POT1 interacted with these non-telomeric binding sites with high affinity and specificity, even when bound to its dimerization partner TPP1. This intrinsic ability of POT1 to recognize NT motifs raises the possibility that the protein may fulfill additional functions at certain non-telomeric locations of the genome, in perhaps gene transcription, replication, or repair.

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