Distinct functions of POT1 at telomeres.
The mammalian protein POT1 binds to telomeric single-stranded DNA (ssDNA), protecting chromosome ends from being detected as sites of DNA damage. POT1 is composed of an N-terminal ssDNA-binding domain and a C-terminal protein interaction domain. With regard to the latter, POT1 heterodimerizes with the protein TPP1 to foster binding to telomeric ssDNA in vitro and binds the telomeric double-stranded-DNA-binding protein TRF2. We sought to determine which of these functions-ssDNA, TPP1, or TRF2 binding-was required to protect chromosome ends from being detected as DNA damage. Using separation-of-function POT1 mutants deficient in one of these three activities, we found that binding to TRF2 is dispensable for protecting telomeres but fosters robust loading of POT1 onto telomeric chromatin. Furthermore, we found that the telomeric ssDNA-binding activity and binding to TPP1 are required in cis for POT1 to protect telomeres. Mechanistically, binding of POT1 to telomeric ssDNA and association with TPP1 inhibit the localization of RPA, which can function as a DNA damage sensor, to telomeres.
Barrientos
Katharine S
KS
Department of Pharmacology and Cancer Biology, Department of Radiation Oncology, Duke University Medical Center, Durham, NC 27710, USA.
Kendellen
Megan F
MF
Freibaum
Brian D
BD
Armbruster
Blaine N
BN
Etheridge
Katherine T
KT
Counter
Christopher M
CM
D002460:Cell Line; D004249:DNA Damage; D004277:DNA, Single-Stranded / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008954:Models, Biological; D050505:Mutant Proteins / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D011485:Protein Binding; D021381:Protein Transport; D051716:Replication Protein A / Q000378:metabolism; D000089804:Shelterin Complex; D016615:Telomere / Q000235:genetics / Q000378:metabolism*; D034501:Telomere-Binding Proteins / Q000378:metabolism*; D035341:Telomeric Repeat Binding Protein 2 / Q000378:metabolism
C495876:ACD protein, human; D004277:DNA, Single-Stranded; D050505:Mutant Proteins; C440849:POT1 protein, human; D051716:Replication Protein A; D000089804:Shelterin Complex; D034501:Telomere-Binding Proteins; D035341:Telomeric Repeat Binding Protein 2