TY - JOUR
T1 - DNA polymerase δ stalls on telomeric lagging strand templates independently from G-quadruplex formation
AU - Lormand, Justin D.
AU - Buncher, Noah
AU - Murphy, Connor T.
AU - Kaur, Parminder
AU - Lee, Marietta Y.
AU - Burgers, Peter
AU - Wang, Hong
AU - Kunkel, Thomas A.
AU - Opresko, Patricia L.
N1 - Funding Information:
NIH [R01ES0515052 to P.L.O., R01GM032431 to P.M.B., R00ES016758 to H.W., R01GM031973 to M.Y.L.] (in part); Division of Intramural Research of the National Institutes of Health, NIEHS [Z01 ES065070 to T.A.K.]. Funding for open access charge: Scaife Foundation through the Center for Nucleic Acids Science and Technology at Carnegie Mellon University (to P.L.O.).
PY - 2013/12
Y1 - 2013/12
N2 - Previous evidence indicates that telomeres resemble common fragile sites and present a challenge for DNA replication. The precise impediments to replication fork progression at telomeric TTAGGG repeats are unknown, but are proposed to include G-quadruplexes (G4) on the G-rich strand. Here we examined DNA synthesis and progression by the replicative DNA polymerase δ/proliferating cell nuclear antigen/replication factor C complex on telomeric templates that mimic the leading C-rich and lagging G-rich strands. Increased polymerase stalling occurred on the G-rich template, compared with the C-rich and nontelomeric templates. Suppression of G4 formation by substituting Li+ for K+ as the cation, or by using templates with 7-deaza-G residues, did not alleviate Pol δ pause sites within the G residues. Furthermore, we provide evidence that G4 folding is less stable on single-stranded circular TTAGGG templates where ends are constrained, compared with linear oligonucleotides. Artificially stabilizing G4 structures on the circular templates with the G4 ligand BRACO-19 inhibited Pol δ progression into the G-rich repeats. Similar results were obtained for yeast and human Pol δ complexes. Our data indicate that G4 formation is not required for polymerase stalling on telomeric lagging strands and suggest that an alternative mechanism, in addition to stable G4s, contributes to replication stalling at telomeres.
AB - Previous evidence indicates that telomeres resemble common fragile sites and present a challenge for DNA replication. The precise impediments to replication fork progression at telomeric TTAGGG repeats are unknown, but are proposed to include G-quadruplexes (G4) on the G-rich strand. Here we examined DNA synthesis and progression by the replicative DNA polymerase δ/proliferating cell nuclear antigen/replication factor C complex on telomeric templates that mimic the leading C-rich and lagging G-rich strands. Increased polymerase stalling occurred on the G-rich template, compared with the C-rich and nontelomeric templates. Suppression of G4 formation by substituting Li+ for K+ as the cation, or by using templates with 7-deaza-G residues, did not alleviate Pol δ pause sites within the G residues. Furthermore, we provide evidence that G4 folding is less stable on single-stranded circular TTAGGG templates where ends are constrained, compared with linear oligonucleotides. Artificially stabilizing G4 structures on the circular templates with the G4 ligand BRACO-19 inhibited Pol δ progression into the G-rich repeats. Similar results were obtained for yeast and human Pol δ complexes. Our data indicate that G4 formation is not required for polymerase stalling on telomeric lagging strands and suggest that an alternative mechanism, in addition to stable G4s, contributes to replication stalling at telomeres.
UR - http://www.scopus.com/inward/record.url?scp=84890376913&partnerID=8YFLogxK
U2 - 10.1093/nar/gkt813
DO - 10.1093/nar/gkt813
M3 - Article
C2 - 24038470
AN - SCOPUS:84890376913
VL - 41
SP - 10323
EP - 10333
JO - Nucleic Acids Research
JF - Nucleic Acids Research
SN - 0305-1048
IS - 22
ER -